Design, synthesis, and SAR of new pyrrole-oxindole progesterone receptor modulators leading to 5-(7-fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)- 1-methyl-1H-pyrrole-2-carbonitrile (WAY-255348)

Article abstract of DOI:10.1021/jm701080t

We have continued to explore the 3,3-dialkyl-5-aryloxindole series of progesterone receptor (PR) modulators looking for new agents to be used in female healthcare: contraception, fibroids, endometriosis, and certain breast cancers. Previously we reported

Full text of DOI:10.1021/jm701080t

J. Med. Chem. 2008, 51, 1861–1873
1861
Design, Synthesis, and SAR of New Pyrrole-Oxindole Progesterone Receptor Modulators
Leading to 5-(7-Fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-methyl-1H-pyrrole-2-
carbonitrile (WAY-255348)
Andrew Fensome,*,† William R. Adams,‡ Andrea L. Adams,‡ Tom J. Berrodin,§ Jeff Cohen,§ Christine Huselton,‡
Arthur Illenberger,4 Jeffrey C. Kern,† Valerie A. Hudak,† Michael A. Marella,† Edward G. Melenski,† Casey C. McComas,†
Cheryl A. Mugford,‡ Ov D. Slayden, Matthew Yudt,§ Zhiming Zhang,§ Puwen Zhang,† Yuan Zhu,§ Richard C. Winneker,§ and
Jay E. Wrobel†
Chemical and Screening Sciences, Biotransformation DiVision, Drug Safety and Metabolism, Women’s Health and Musculoskeletal Biology,
and BioResources, Wyeth Research, 500 Arcola Road, CollegeVille, PennsylVania 19426, and DiVision of ReproductiVe Sciences, Oregon
National Primate Research Center, Oregon Health and Science UniVersity, BeaVerton, Oregon 97005
ReceiVed August 31, 2007
We have continued to explore the 3,3-dialkyl-5-aryloxindole series of progesterone receptor (PR) modulators
looking for new agents to be used in female healthcare: contraception, fibroids, endometriosis, and certain
breast cancers. Previously we reported that subtle structural changes with this and related templates produced
functional switches between agonist and antagonist properties (Fensome et al. Biorg. Med. Chem. Lett.
2002, 12, 3487; 2003, 13, 1317). We herein report a new functional switch within the 5-(2-oxoindolin-5-
yl)-1H-pyrrole-2-carbonitrile class of compounds. We found that the size of the 3,3-dialkyl substituent is
important for controlling the functional response; thus small groups (dimethyl) afford potent PR antagonists,
whereas larger groups (spirocyclohexyl) are PR agonists. The product from our optimization activities in
cell-based systems and also for kinetic properties in rodents and nonhuman primates was 5-(7-fluoro-3,3-
dimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-methyl-1H-pyrrole-2-carbonitrile 27 (WAY-255348), which
demonstrated potent and robust activity on PR antagonist and contraceptive end points in the rat and also
in cynomolgus and rhesus monkeys including ovulation inhibition, menses induction, and reproductive tract
morphology.
Introduction
The control and regulation of progesterone (1, Figure 1) and
the progesterone receptor (PR)a has been a well-recognized
drug target for female reproductive health for the past four
decades. The PR is a member of the nuclear transcription factor
superfamily.1 PR agonists have been used in contraception
(either alone or in combination with an estrogen), in postmeno-
pausal hormone therapy in combination with estrogens, and also
in high dose treatments for endometriosis and uterine fibroids.
Antagonists have been studied clinically for contraception,2,3
for the treatment of uterine fibroids,4–8 and for the termination
of pregnancy.9 Ligands for this receptor display a spectrum of
activities from full agonist (e.g., progesterone 1 and medrox-
Figure 1. Steroidal PR modulators: progesterone 1, medroxyproges-
yprogesterone acetate 2) to antagonist (e.g., mifepristone 3 and
terone acetate 2, mifepristone 3, 4, asoprisnil 5.
ORG-3362810 4). Newer compounds, such as asoprisnil11 5, fall
somewhere in between and are described as selective proges-
and estrogen receptor (ER)), which can result in unwanted side
terone receptor modulators (SPRM). What is notable about the
effects, which potentially limit the benefits of the desired
clinically studied PR ligands is the common steroidal platform.
therapy. Analogous to the estrogen receptor field, where great
This platform brings with it potential to cross-react with other
advances have been made with nonsteroidal modulators,12–15
members of the steroid family (androgen receptor (AR),
more recent work in the PR modulator field has been applied
glucocorticoid receptor (GR), mineralocorticoid receptor (MR),
to nonsteroidal templates.16,17
Over the past few years, we have published new PR agonist
templates such as the thio-oxindoles18 6 and cyanopyrrole
* Telephone 484-865-5515, fax 484-865-9398, e-mail fensoma@
wyeth.com.
†
carbamates 7,19 and the synergistic combination of both features
(tanaproget 8),20 as well as PR antagonists such as the oxindoles
9 and 10.21 What has become apparent during the course of
this work is the subtle structure–activity relationship (SAR) that
controls the functional outcome from agonist to antagonist,
without relying upon the larger structural changes necessary for
their steroidal counterparts (e.g., the 11-ꢀ-aryl moiety present
in 3–5). In this paper, we describe a recent SAR exploration of
Chemical and Screening Sciences, Wyeth Research.
Drug Safety and Metabolism, Wyeth Research.
Women’s Health and Musculoskeletal Biology, Wyeth Research.
BioResources, Wyeth Research.
‡
§
4
Oregon Health and Science University.
a Abbreviations: PR, progesterone receptor; AR, androgen receptor; GR,
glucocorticoid receptor; MR, mineralocorticoid receptor; ER, estrogen
receptor; RLM, CMLM, and HLM, rat, cynomolgus monkey, and human
liver microsomes, respectively.
10.1021/jm701080t CCC: $40.75
2008 American Chemical Society
Published on Web 03/05/2008

1862 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Fensome et al.
catalyzed ring closure (p-TsOH in EtOH) to afford the fluori-
nated oxindoles 34a-c. Alkylation of the 3-position was
effected by generation of the dianion under the action of
n-BuLi-LiCl in THF, followed by reaction with either an alkyl
iodide to form the intermediates for targets 25–28 or an alkyl
di-iodide for the spirocycles 29 and 30. Compounds 35a-f were
then brominated under standard conditions to afford coupling
partners 36a-f. The 4-, 6-, and 7-fluoro-3,3-dimethyl targets
25, 26, and 27 were initially prepared via coupling with boronic
acid 16 under traditional coupling conditions (Pd(Ph3P)4, K2CO3,
THF, reflux); however the yields were low (5–28%).
Compound 27 was selected for optimization of the coupling
chemistry between the bromide 36c and boronic acid 16, Table
1. We selected tris-dibenzylideneacetone dipalladium(II) as the
metal source, and a range of phosphines were surveyed. KF
was used as the base. Reactions were poor with the nonhindered
phosphines (entries 1 and 2); however the bulkier phosphines,
such as dicyclohexyl phenylphosphine (entry 3) and Q-phos
(entry 4), gave more promising results. We observed a solvent
effect and found that THF and dioxane gave better yields than
did toluene (entry 4 and 5). Following the work of Fu and co-
workers,24 we then examined tri-tert-butylphosphine as the
ligand (entries 6–8) and found the stoichiometry between the
Pd2(dba)3 and the phosphine to be important; the optimal
conditions found were a ratio of 1:2 between the Pd2-
(dba)3 ·CHCl3 and the tri-tert-butylphosphine in THF giving 27
in an isolated yield of 84%. Under these optimized conditions,
the diethyl derivative 28 and the 3,3-spirocycloalkyl compounds
29 and 30 were prepared from their corresponding bromides
36a-c.
Figure 2. Nonsteroidal PR modulators.
Table 1. Optimization of Coupling Reaction To Prepare 27 from
Bromide 36c and Boronic Acid 16
ratio
yield
(%)
catalyst
Pd(Ph3P)4
Pd2(dba)3 ·CHCl3 (o-tol)3P
Pd2(dba)3 ·CHCl3 (C6H11)2PPh
Pd2(dba)3 ·CHCl3 Q-phosa
Pd2(dba)3 ·CHCl3 Q-phosa
Pd2(dba)3 ·CHCl3 (t-Bu)3P
ligand
Pd/P
base
solvent
1
2
3
4
5
6
7
8
K2CO3 DMF
19
0
KF
KF
KF
KF
dioxane
dioxane
dioxane
toluene
dioxane
dioxane
THF
58
64
30
30
28
80
1:3 KF
Pd2(dba)3 ·CHCl3 Pd((t-Bu)3P)2 1:3 KF
Pd2(dba)3 ·CHCl3 (t-Bu)3P 1:2 KF
a Q-phos ) di-tert-butylphosphinopentaphenylferrocene.
a series of pyrrole-oxindoles 11 (Figure 2), their synthesis, and
their biological activities. The work described herein led to the
potent orally active PR antagonist 5-(7-fluoro-3,3-dimethyl-2-
oxo-2,3-dihydro-1H-indol-5-yl)-1-methyl-1H-pyrrole-2- carbo-
nitrile 27 (WAY-255348).22
Chemistry
We then further sought to expand the oxindole template by
incorporating functionality on N-1 through alkylation of 27 to
afford compounds 37–48, Scheme 4. Treating 27 with potassium
tert-butoxide in THF and reacting the anion with the respective
electrophile afforded the desired products.
The 3,3-unsubstituted oxindole derivatives 12 and 13 were
prepared according to Scheme 1. Palladium-catalyzed coupling
of the bromide 14 with N-Boc-pyrrole-2-boronic acid gave the
pyrrole 15. Installation of the nitrile to provide target compound
12 was accomplished by treatment with chlorosulfonyl isocy-
anate followed by N,N-dimethylformamide (DMF) quench, and
subsequent Boc group removal was achieved by thermolysis in
dimethylacetamide (DMA). The analogous N-methyl derivative
13 was prepared via a palladium-catalyzed coupling of 14 with
the boronic acid 16, which was prepared by treatment of the
pyrrole 17 with lithium di-isopropyl amide (LDA) and tri-
isopropyl borate in tetrahydrofuran (THF).
Results and Discussion
The compounds synthesized were tested for functional activity
in the human T47D cell alkaline phosphatase assay.25 In the
agonist mode, stimulation of alkaline phosphatase is reported
as an EC50 value. In the antagonist mode, the compound is tested
for its ability to antagonize the alkaline phosphatase induced
by progesterone (1 nM) and is reported as an IC50. Compounds
were then assayed in a T47D whole cell competition-binding
assay displacing tritiated R5020, the activity being reported as
an IC50.
The SAR in the oxindole series is described below, Table 2.
The unsubstituted oxindole 12 was a functional antagonist in
the T47D alkaline phosphatase assay (IC50 ) 300 nM). Addition
of a methyl group to the pyrrole nitrogen afforded a 10-fold
increase in activity (13, 20 nM). Further addition of substituents
to the 3-position of the oxindole again increased potency; for
example, the 3,3-dimethyl derivative (18, IC50 ) 10 nM) and
the 3,3-diethyl compound (19, IC50 ) 3.3 nM) were also potent
PR antagonists. We then chose to prepare the larger spirocyclic
derivatives. The spirocyclopentyl derivative 20 was found to
have a biphasic curve in the T47D alkaline phosphatase assay
with approximate IC50 ) 30 nM and EC50 ) 300 nM. The next
higher homologue, the spirocyclohexyl derivative 21, was found
to be a potent and efficacious agonist (T47D EC50 ) 2.3 nM).
It is interesting to compare the cyano-pyrrole series with the
previously reported 5-aryl derivatives, the dimethyl derivative
9 and spirocyclohexyl derivative 10, where the compounds were
functional antagonists regardless of the size of the 3,3-alkyl
The des-fluoro 3,3-dialkylated compounds 18–21 were pre-
pared following Scheme 2.
Oxindole 22 was treated with 2 equiv of n-BuLi in THF at
-25 °C in the presence of N,N,N′,N′-tetramethylethylenediamine
(N,N,N′,N′-TMEDA) to generate the dianion, which was sub-
sequently quenched with either an alkyl iodide to prepare the
intermediates 23a,b for targets 18 and 19 or an alkyl di-iodide
for the preparation of the intermediates 23c,d needed for the
spirocyclic targets 20 and 21.23 The products were then
brominated under standard conditions (Br2, NaOAc, AcOH) to
give the 5-bromo oxindoles 24a-d respectively. The bromides
24a-d were then coupled with the pyrrole boronic acid 16,
either in the isolated state or generated in situ from the treatment
of pyrrole 17 with LDA and tri-isopropyl borate, to provide
the target compounds 18–21.
The fluorinated analogs 25–30 of general target 11 were
prepared via a malonate route, Scheme 3.
Reaction of the 2-fluoro-nitrobenzenes 31a-c with dimethyl
malonate in DMF in the presence of potassium carbonate
afforded malonates 32a-c, which were subsequently hydrolyzed
and decarboxylated in refluxing 6 N HCl to afford the acetic
acids 33a-c. Hydrogenolysis over Pd-C was followed by acid-

Pyrrole-Oxindole Progesterone Receptor Modulators
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1863
Scheme 1a
a Reagents and conditions: (a) N-Boc-pyrrole-2-boronic acid, Na2CO3, Pd(Ph3P)4, THF, reflux; (b) (i) ClSO2NCO, DCM, -78 °C, (ii) DMF, rt; (c) DMA,
175 °C; (d) LDA, (iPrO)3B, THF, -78 °C; (e) Pd(Ph3P)4, K2CO3, 16, THF, reflux.
Scheme 2a
18, 25, and 27, respectively). Similarly in the whole-cell binding
assay, a small increase in potency was noted for both compounds
25 and 27 (IC50 ) 5.5 and 4.1 nM, respectively). In comparison
with mifepristone 3, 27 was approximately 10-fold less active
on both T47D alkaline phosphatase and whole cell binding
assays. In contrast, the 6-fluoro derivative 26 was slightly less
active than the parent 18 (T47D alkaline phosphatase IC50
)
20 nM). In the microsome stability assay, the 4-fluoro derivative
25 lost RLM stability in comparison with the des-fluoro
homologue 18 (RLM t1/2 ) 2 and 6 min, respectively, for 18
and 25). The 6- and 7-fluoro derivatives 26 and 27 did show
significant improvements in rat liver microsome data over
compound 18 (RLM t1/2 ) 30 and 26 min, respectively, for 26
and 27). Compound 27 showed a small improvement in
cynomolgus monkey microsome stability over the des-fluoro
homologue 18.
a Reagents and conditions: (a) n-BuLi, N,N,N′,N′-TMEDA, THF, -25
°C, alkyl iodide; (b) Br2, NaOAc, AcOH, rt; (c) Pd(Ph3P)4, K2CO3, 16,
THF, reflux.
The 7-fluoro-3,3-diethyl derivative 28 had similar functional
potency when compared with its des-fluoro congener 19 (T47D
IC50 ) 4.9 and 3.3 nM for 28 and 19, respectively). Addition
of the 7-fluorine substituent to the spirocyclopentyl derivative
20 shifted the mixed agonist/antagonist functional activity in
favor of the antagonist mode (IC50 ) 3.2 nM, 85% inhibition,
compound 29), while the 7-fluoro-3,3-spirocyclohexyl derivative
30 remained a relatively potent agonist (EC50 ) 1.3 nM).
To further capitalize upon the pyrrole-oxindole series, we
selected compound 27 as a platform from which to explore the
nature of the oxindole nitrogen substituent, Table 5. In general,
small substituents gave the most potent antagonists in the T47D
alkaline phosphatase assay. Thus the methyl derivative 37 gave
comparable functional activity to the N-H congener 27 (IC50
) 7.4 nM). The ethyl, n-propyl, and iso-butyl derivatives all
lost between 2- and 4-fold in potency (IC50 ) 29, 27, 13, and
30 nM for compounds 38, 39, 40, and 41, respectively). A size
limitation was observed in the cycloalkyl compounds, where
the cyclopentane 42 was similar in activity to the smaller alkyl
substituents but the cyclohexyl derivative 43 lost approximately
substituent (T47D IC50 ) 27 and 14.7 nM for 9 and 10,
respectively).21 In comparison with mifepristone 3, the most
potent of the unsubsitituted oxindole derivatives, the diethyl
analog 19 was approximately 1 order of magnitude less active
in the T47D functional activity.
In the whole cell-binding assay the unsubstituted oxindole
13 was a competitive ligand for the hPR (IC50 ) 80 nM).
Similarly, compounds 18 and 19 were also competitive (IC50
) 22 and 35 nM, respectively) but gained potency due to the
3,3-dialkyl substitution, commensurate with the increase in
potency seen in the functional alkaline phosphatase assay. The
agonist 21 displayed potent displacement of the radioligand (IC50
) 3.3 nM), in close agreement with its functional activity. In
the same whole cell binding assay, mifepristone 3 was ap-
proximately 2 orders of magnitude more potent than the oxindole
antagonists (T47D whole cell binding assay IC50 ) 0.6 nM).
As part of the characterization of the compounds, female rat,
human, and cynomolgus monkey microsome stability data was
obtained for a subset of compounds, Table 3.
While compound 13 was found to be stable across all three
species tested, the 3,3-dimethyl 18 and 3,3-diethyl 19 derivatives
were found to have low stability in the rat liver microsomes
and moderate stability in cynomolgus monkey microsomes. No
metabolic stability issues were noted for any of the compounds
tested in human liver microsomes.
Controlling metabolic stability of compounds, either by
blocking a site of metabolism or by reducing the electron density
of a ring system through induction, can affect pharmacokinetic
properties.26 We selected the dimethyl oxindole 18 to conduct
our studies and selectively installed a fluorine atom at the 4-,
6-, and 7-positions of the benzo-fused ring system, Table 4.
Addition of fluorine to either the 4- or 7-positions of the
dimethyl derivative 18 afforded a small increase in potency in
the alkaline phosphatase assay (IC50 ) 10, 5.9, and 5.0 nM for
40-fold in potency relative to the N-H derivative 27 (IC50
)
25 and 194 nM for 42 and 43, respectively). We found that
this series of molecules would also tolerate unsaturation, thus
the allyl 44 and propargyl 45 derivatives were similar in potency
to the smaller alkyl compounds (T47D alkaline phosphatase IC50
) 4.5 and 30 nM, respectively). Larger aromatic functionalities
(benzyl 46 and phenethyl 47) were only poorly tolerated. We
also prepared one more functionalized derivative, the acetate
48, which had modest potency in the functional activity assay
(IC50 ) 48 nM).
To further profile 27, the ability of the compound to cross-
react with other members of the steroid receptor family was
evaluated in a Gal4 mammalian one-hybrid assay,27 Table 6.

1864 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Fensome et al.
Scheme 3a
a Reagents and conditions: (a) K2CO3, CH2(CO2Me)2, DMF, 65 °C; (b) 6 N HCl, reflux; (c) (i) H2, Pd-C, AcOH, (ii) p-TsOH, EtOH, reflux; (d) n-BuLi,
LiCl, THF, -78 °C, then alkyl iodide; (e) Br2, CH2Cl2-AcOH; (f) Pd(Ph3P)4, K2CO3, 16, THF, reflux for 25–27 or (g) Pd2(dba)3 ·CHCl3, t-Bu3P, KF, 16,
THF, rt for 27–30.
Scheme 4a
Table 3. Rat, Cynomolgus Monkey and Human Liver Microsome
Stability Data for 13, 18–26, and 27
compound RLMa t1/2 (min) CMLMb t1/2 (min) HLMc t1/2 (min)
13
18
19
25
26
27
>30
6
10
2
30
26
>30
13
15
d
d
19
>30
>30
>30
d
d
>30
a Rat liver microsome. b Cynomolgus monkey liver microsome. c Human
liver microsome. d Not determined.
a Reagents and conditions: (a) t-BuOK, THF, electrophile.
mammalian one-hybrid assay (IC50 ) 6.9, 6.1, and 196 nM,
respectively, for 3, 5, and 27). However in terms of the PR/AR
selectivity ratio, 27 and 3 are equivalent (PR/AR ) 39- and
34-fold, respectively, for 27 and 3). In the glucocorticoid
receptor (GR) assay, the well-characterized antagonist activity
of 3 was apparent (IC50 ) 0.6 nM); asoprisnil 5 was somewhat
less potent (IC50 ) 85 nM), whereas 27 was inactive as either
an agonist or antagonist of this receptor, a characteristic not
shared by the steroidal PR antagonists. Against both the
mineralocortocoid (MR) and estrogen receptors (ER), 27 had
Table 2. Alkaline Phosphatase and Whole Cell Binding Data in T47D
Cells for Compounds 3, 6–10, 12, 13, and 18–21
T47D whole-cell
T47D alk phosa
binding
no agonist activity and only weak antagonist activity (IC50
1600 and 1900 nM for hMR and hER, respectively).
)
IC50
R3 (nM)
EC50
(nM)
IC50
(nM)
compd
R1
R2
In ViWo Pharmacology. Compounds 18 from the des-fluoro
series and 27 from the 7-fluoro series were chosen for further
in ViVo characterization. These compounds were selected based
upon their similar potency in the T47D alkaline phosphatase
assay and to examine the effect of the 7-fluorine present in 27.
Pharmacokinetic parameters for both compounds 18 and 27
were obtained in female Sprague–Dawley rats, Table 7.
Compounds 18 and 27 were dosed in separate 1 in 4 iv
cassette studies in female rats. It can be seen from the data that
27 in comparison to 18 achieves a higher exposure (AUC0-inf
) 937 and 1978 ng ·h/mL for 18 and 27, respectively) with a
longer half-life (t1/2 ) 3.0 and 9.5 h for 18 and 27, respectively).
These effects are probably due to the lower clearance for the
latter compound, which is in agreement with what would be
expected from the increased rat metabolic stability observed for
27. The discrete iv study for 27 was in close agreement with
3
6
7
8
0.2
0.6
b
4.9
0.5
b
2.0
1.1
0.15
9
27
15
300
20
10
3.3
10
12
13
18
19
20
21
25
b
H
H
Me
Et
H
H
Me
Et
H
Me
Me
Me
80
22
36
8.1
3.3
-CH2CH2CH2CH2-
-CH2CH2CH2CH2CH2- Me
Me ∼30 ∼300
2.3
a T47D cell alkaline phosphatase functional activity assay. Values
represent the average of at least duplicate determinations. The standard
deviation for the assay was typically (20% of mean or less. b Not
determined.
In comparison with mifepristone 3 and asoprisnil 5, 27 is a
relatively weak androgen receptor (AR) antagonist in the Gal4

Pyrrole-Oxindole Progesterone Receptor Modulators
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1865
Table 4. Alkaline Phosphatase and Whole Cell Binding Data in T47D Cells for Compounds 3 and 25–30
T47D alk phosa
T47D whole cell binding
IC50 (nM)
compd
R1
R2
R3
R4
R5
IC50 (nM)
EC50 (nM)
3
0.2
5.9
20.3
5.0
0.6
5.5
b
4.1
b
25
26
27
28
29
30
Me
Me
Me
Et
Me
Me
Me
Et
F
H
F
H
H
H
H
H
H
F
F
F
F
H
H
H
H
H
4.9
-CH2CH2CH2CH2-
-CH2CH2CH2CH2CH2-
3.2 (85%)c
∼3000
14.7
3.6
1.3
a T47D cell alkaline phosphatase functional activity assay. Values represent the average of at least duplicate determinations. The standard deviation for
the assay was typically (20% of mean or less. b Not determined. c Inhibition (percent) relative to R5020 as reference.
Table 5. Alkaline Phosphatase T47D Cell Data for Compounds 27 and
37–47
to antagonize the progesterone-induced decidualization response
in the stromal cells of the uterus.28 In this assay, both compounds
dose-dependently inhibited the decidual response produced by
progesterone following oral administration in an aqueous vehicle
(0.5% methyl cellulose/2% Tween 80) with potencies compa-
rable to the reference compound mifepristone 3 (ED50 ) 0.2,
0.3, and 0.7 mg/kg for 18, 27, and 3, respectively).
To establish efficacy markers for pursuing contraception as
our primary clinical indication, we examined both compounds
18 and 27 in the menses induction assay in female cynomolgus
monkeys. A successful outcome in this assay was used as a
screen prior to establishing efficacy in the rhesus macaque
ovulation inhibition and reproductive tract model.
In mature normal cycling female cynomolgus monkeys,
administration of a PR antagonist, mifepristone 3, during the
midluteal phase (days 19–22 of the menstrual cycle) causes
premature menstruation indicating PR antagonist activity.29 The
clinical relevance of menses induction to a contraceptive
regimen has been reported for 4 and a second steroidal
antagonist 11-[4-(dimethylamino)phenyl]-4′,5′-dihydro-6-meth-
yl-(6ꢀ,11ꢀ,17ꢀ)-spiro[estra-4,9-diene-17,2′(3′H)-furan]-3-one
(ORG-31710), where administration of a single dose of this
compound to women taking the 75 µg desogestrel progestin
only pill during the luteal phase of the menstrual cycle was
effective at inducing early menstruation.30
compound
R
T47D alk phosa IC50 (nM)
3
0.2
5
15
29
27
13
30
25
194
4.5
30
164
351
48
27
37
38
39
40
41
42
43
44
45
46
47
48
H
Me
Et
n-Pr
i-Pr
i-Bu
cyclopentyl
cyclohexyl
allyl
prop-2-ynyl
benzyl
2-phenethyl
methylacetate
a T47D cell alkaline phosphatase functional activity assay. Values
represent the average of at least duplicate determinations. The standard
deviation for the assay was typically ( 20% of mean or less.
Table 6. Steroid Receptor Cross Reactivity Data for Compounds 3, 5,
and 27a
A typical menstrual cycle in cynomolgus monkeys is between
28 and 33 days in length. In an animal treated with an active
PR antagonist for 4 days starting on day 19–22 of the menstrual
cycle, the cycle length is reduced to 22–24 days. Steroidal PR
antagonists mifepristone 3 and 4, in our own hands, dosed at 5
mg/kg orally, induced early menses in 60% (3 out of 5 animals)
and 50% (2 out of 4 animals), respectively, Table 8. Compound
18 following an oral dose of 5 mg/kg was inactive (0 out of 2
animals). In contrast, compound 27 at 5 mg/kg po induced early
menses in 75% (3 out of 4 animals) of cycling monkeys.
To help understand the difference in activities seen in the
menses induction assay between compounds 18 and 27, the
female cynomolgus monkey pharmacokinetic parameters were
measured, Table 9.
hARb
EC50 IC50
compd (nM) (nM) (nM) (nM) (nM) (nM) (nM) (nM)
hGRc
hMRd
hERe
EC50 IC50 EC50 IC50 EC50 IC50
3
5
27
f
f
f
6.9
6.1
196
f
f
f
0.6
85
f
g
f
f
g
1600
3700
g
f
f
g
1900
f
a Steroid receptor cross-reactivity determined in a Gal4 mammalian
hybrid assay. Values represent the average of at least duplicate determina-
tions. The standard deviation for the assay was typically (20% of mean or
less. b hAR ) human androgen receptor; antagonist mode run in the presence
of 5R-dihydrotestosterone (10 nM). c hGR ) human glucocorticoid receptor;
antagonist mode run in the presence of dexamethasone (5 nM). d hMR )
human mineralocorticoid receptor; antagonist mode in the presence of
aldosterone (3 nM). e hER ) human estrogen receptor; antagonist mode
run in the presence of 17ꢀ-estradiol (5 nM). f Not statistically active. g Not
determined.
In the monkey dosed at 0.25 mg/kg iv (1 in 4 cassette dosed
in the same study), compound 27 had a longer half-life (t1/2
)
the data obtained from the cassette study, allowing for differ-
ences in the dose administered. Following oral dosing, 27
showed good exposure (AUC0-inf ) 7149 ng·h/mL), a long
half-life (t1/2 ) 9.3 h), and moderate bioavailability (F ) 31%).
Compounds 18 and 27 were evaluated in the female rat
decidual model. This model measures the ability of a compound
8.8 and 2.1 h for 27 and 18, respectively), lower clearance,
higher volume of distribution, and higher exposure than did des-
fluoro analog compound 18. Dosed orally at 5 mg/kg, compound
27 had a good exposure (AUC0-inf ) 5738 ng·h/mL) and good
bioavailability (F ) 49%) in this species. Although both
compounds have similar potency in the T47D cell alkaline

1866 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Table 7. Pharmacokinetic Properties for Compounds 18 and 27a
Fensome et al.
compound study designb route dose (mg/kg) C0 (ng/mL) Cmax (ng/mL) t1/2 (h) Clp (mL/(min ·kg)) Vss (L/kg) AUC0-inf (ng·h/mL) %F
18
27
1 in 4
1 in 4
discrete
discrete
ivc
iv
0.5
0.5
1
288
385
528
3.0
9.5
9.4
9.3
9
4
4
2.2
2.9
2.4
937
1978
4650
7149
iv
pod
5
414
31
a Pharmacokinetic properties generated in female Sprague–Dawley rats. b Compounds were dosed as 1 in 4 cassette studies or in discrete studies. c Compounds
dosed iv in DMSO/PEG200 20:80 vehicle. d Compounds dosed po in aqueous 2% Tween 80/0.5% methyl cellulose vehicle.
Table 8. Cynomolgus Monkey Menses Induction Data for 3, 4, 18, and
were collected as needed to identify the midcycle surge of
estradiol. Ovulation was monitored by blood ovarian steroid
profile. The reproductive tract was collected at the end of study
for histological and immunocytochemical analyses.
27a
compound
3
4
18
27
animals with early menstruation (%)
60
50
0
75
a Compounds dosed orally at 5 mg/(kg ·day) on days 19–22 of the
menstrual cycle. A positive result was given if menses occurred on days
22–24. Compounds were dosed in aqueous 2% Tween 80/0.5% methyl
cellulose vehicle.
Similar to the cynomolgus study above none of the animals
treated with 27 showed a rise in luteal phase progesterone,
indicating that ovulation was blocked. Only one animal in this
group showed a significant midcycle estradiol surge. Laparo-
scopic examination revealed no evidence of an ovulatory site
on the ovaries of this animal. No laparoscopic examination was
conducted on the other two animals because there was no
evidence of an estradiol surge or increase in serum progesterone
indicative of a functional corpus luteum.
phosphatase assay and also in the rat decidualization assay, the
lack of activity for compound 18 in the cynomolgus monkey
menses induction assay and positive response for compound
27 can probably be attributed to the pharmacokinetic differences
between the two compounds in the primate.
The effect of 27 on the uterus was evaluated at necropsy (day
28), Table 10. All the animals treated with 27 had thin-atrophied
endometria relative to the endometrium from animals collected
during the normal luteal phase (2.2 ( 0.33 mm and 60 ( 21
mg for 27 treated animals; 4.6 ( 0.7 mm and 430 ( 26 mg for
luteal-phase vehicle control). Compared with luteal phase
animals, which have secretory endometria with sacculated
glands, the endometrium of 27 treated animals was generally
proliferative in appearance with tubular glands. Myometrial
compaction is also apparent for 27 treated animals (1760 ( 517
mg and 2213 ( 177 mg for 27 and luteal-phase control animals,
respectively).
The ability of PR antagonists to block ovulation in humans
has been demonstrated with mifepristone and with other
steroids.31 At higher doses, inhibition of follicular development
occurs by suppressing the gonadotropin surge. At lower doses
however, a delay in ovulation is induced rather than complete
inhibition, which is also accompanied by a disruption of
endometrial development.32 Our nonhuman primate ovulation
inhibition assay was based upon the model published by Hodgen
and co-workers, who demonstrated that mifepristone 3 inhibited
ovulation in 5 out of 6 normally cycling cynomolgus monkeys
(1 mg/kg im).33
To determine the effect of compound 27 on ovulation in the
nonhuman primate, normal cycling cynomolgus monkeys
were treated orally with 27 for 30 days starting on day 2 of the
menstrual cycle (day 1 ) first day of menses), Figure 3. The
compound (10, 3, or 1 mg/kg) was given daily in 2% Tween
80/0.5% methylcellulose vehicle. Blood samples were taken 3
times per week (Monday, Wednesday, and Friday), and ovula-
tion was monitored through the measurement of serum proges-
terone levels. Animals treated with vehicle showed elevated
progesterone levels during luteal phase, indicating the formation
of a functional corpus luteum after ovulation. In contrast, all
four animals treated with 10 mg/kg of 27 showed only basal
levels of progesterone throughout the entire treatment cycle,
suggesting complete inhibition of ovulation. Among the six
animals treated with 3 mg/kg of 27, only one monkey had a
luteal phase progesterone surge. The other five animals showed
only basal levels of progesterone, indicating that no ovulation
occurred. Among the four monkeys treated with 1 mg/kg of
27, one monkey showed inhibition of ovulation. Ordinary least-
squares analysis of this data indicates an ED50 for ovulation
inhibition of approximately 1.6 mg/kg.
The menses induction, ovulation inhibition, and reproductive
tract histology are consistent with compound 27 behaving as a
progesterone antagonist in the primate.
In summary, we have further explored the SAR in the 5-aryl-
oxindole class of progesterone receptor modulators. We found
a subseries of cyano-pyrroles that possess an additional func-
tional motif, the size of the 3,3-substituent, which determines
the functional activity (agonist or antagonist) of the compound
in Vitro. Selected examples were found that have excellent in
ViVo properties in the rat decidualization model, possessing
potencies similar to the steroidal comparator mifepristone 3.
The pharmacokinetic properties of these compounds support the
observed in ViVo activity. Compound 27 was found to be an
active PR antagonist in the cynomolgus monkey and in the
rhesus macaque. Compound 27 was advanced into phase 0
studies to be studied as a contraceptive agent in women.
Experimental Procedures
Chemistry. Solvents were purchased as anhydrous grade and
used without further purification. Reagents were purchased from
commercial suppliers and used without purification. 1H NMR
spectra were recorded on a Varian Inova 400 instrument with
chemical shifts reported in d values (parts per million, ppm) relative
to an internal standard (tetramethylsilane). High-resolution mass
spectra were obtained on an Agilent TOF. Low-resolution electro-
spray (ESI) mass spectra were recorded on a Waters Alliance ZMD
instrument. Low-resolution electron impact (EI, EE ) 70 eV) mass
spectra were recorded on a Finnigan Trace mass spectrometer.
Analytical thin layer chroimatography (TLC) were performed on
precoated glass plates (silica gel F-254) and visualized under UV
light.
Compound 27 was further evaluated in the rhesus macaque
to study its effect upon ovulation and upon the reproductive
tract. For this study, three adult female rhesus macaques with
regular menstrual cycles were assigned and allowed to complete
one full menstrual cycle. Beginning on day 2 of the next cycle
(day 1 ) first day of menses), the animals were treated with 27
(10 mg/(kg·day), po, 2% Tween 80/0.5% methylcellulose with
artificial cherry flavoring) for 28 days. Blood was collected every
3 days (mid-day), and serum was assayed for estradiol and
progesterone to monitor ovarian function. Additional samples

Pyrrole-Oxindole Progesterone Receptor Modulators
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1867
Table 9. Female Cynomolgus Monkey Pharmacokinetic Parameters for Compounds 18 and 27a
compound study designb route dose (mg/kg) C0 (ng/mL) Cmax (ng/mL) t1/2 (hr) Clp (mL/(min ·kg)) Vss (L/kg) AUC0-inf (ng·h/mL) %F
18
27
1 in 4
1 in 4
discrete
discrete
ivc
ivc
ivc
pod
0.25
0.25
1
241
178
856
2.1
8.8
7.8
12
7
7
1.6
3.9
3.4
343
578
2358
5738
5
179
15.7
49
a Pharmacokinetic properties generated in female cynomolgus monkeys. b Compounds were dosed as 1 in 4 cassette studies or in discrete studies. c Compounds
dosed iv in DMSO/PEG200 20:80 vehicle. d Compounds dosed po in aqueous 2% Tween 80/0.5% methyl cellulose vehicle.
7.15-7.13 (m, 1H), 6.80 (d, J ) 7.9 Hz, 1H), 6.32 (d, J ) 3.8 Hz,
1H), 3.45 (s, 2H), and 1.30 (s, 9H). MS (ESI, [M + H]+) 324.0.
HRMS calculated for C18H17N3O3 + H+ 324.1343; found (ESI,
[M + H]+) 324.1347.
Step 3. 5-(2-Oxoindolin-5-yl)-1H-pyrrole-2-carbonitrile (12).
A solution of tert-butyl 2-cyano-5-(2-oxoindolin-5-yl)-1H-pyrrole-
1-carboxylate (0.23 g, 0.71 mmol) in dry DMA (5 mL) was heated
to 175 °C under nitrogen (30 min). After cooling to room
temperature, the mixture was poured into water and extracted with
EtOAc (×2), washed with water (×2), dried (MgSO4), and
evaporated. The residue was then purified by silica gel column
chromatography (hexanes/acetone 100:0-0:100) to afford the title
1
Figure 3. Ovulation inhibition data for 27 in cycling cynomolgus
monkeys. Ovulation is determined by a luteal phase progesterone surge.
compound 12 (0.80 g, 49%). H NMR (400 MHz, DMSO-d6) δ
12.46 (s, 1H), 10.49 (s, 1H), 7.59 (s, 1H), 7.55 (dd, J ) 8.1 and
The number of animals in each treatment group and those experiencing
1.8 Hz, 1H), 6.95 (d, J ) 3.8 Hz, 1H), 6.85 (d, J ) 8.1 Hz, 1H),
ovulation are indicated above the bars.
6.57 (d, J ) 3.8 Hz, 1H), and 3.53 (s, 2H). HRMS calculated for
C13H10N3O + H+ 224.0828; found (ESI, [M + H]+) 224.0818.
Table 10. Effect of 27 on Ovulation, Uterine and Oviductal
Morphometrics, and Cell Proliferation in Naturally Cycling Macaquesa
1-Methyl-5-(2-oxo-2,3-dihydro-1H-indol-5-yl)-1H-pyrrole-2-
carbonitrile (13). 1-Methyl-1H-pyrrole-2-carbonitrile (0.5 g, 4.8
mmol) and tri-iso-propylborate (1.1 mL, 4.8 mmol) were dissolved
in THF (12 mL) and cooled to 0 °C. LDA (2.5 mL, 5 mmol, 2 M)
was added over 10 min. The reaction was stirred (30 min) and then
allowed to warm up to room temperature.
27 (n ) 3) luteal-phase control (n ) 5)
ovulation rate
0/3
60 ( 21
5/5
endometrial weight (mg)b
430 ( 26
4.6 ( 0.7
2213 ( 177
0
endometrial thickness (mm) 2.2 ( 0.33
In a separate flask, 5-bromoindolin-2-one (0.30 g, 1.42 mmol)
and tetrakis(triphenylphosphine)palladium(0) (0.030 g) were dis-
solved in THF (12 mL) and stirred under nitrogen (15 min). The
above prepared boron complex was transferred (via syringe) to this
solution, followed by addition of potassium carbonate (0.70 g, 5
mmol) and water (6 mL). The mixture was then heated under reflux
(3 h). The reaction mixture was cooled to room temperature, then
poured into water and extracted into EtOAc, dried (MgSO4), and
evaporated. The residue was then purified by silica gel column
chromatography (hexanes/EtOAc 8/2 then 6/4) to afford the title
compound (13) (0.035 g, 11%). 1H NMR (400 MHz, DMSO-d6) δ
10.53 (s, 1H), 7.35 (s, 1H) 7.32 -7.29 (m, 1H), 7.00 (d, J ) 4.0
Hz, 1H), 6.91 (d, J ) 8.0 Hz, 1H), 6.25 (d, J ) 4.0 Hz, 1H), 3.69
(s, 3H), and 3.54 (s, 2H). HRMS calculated for C14H11N3O + H+
238.09749; found (ESI, [M + H]+) 238.0985.
myometrial weight (mg)
functionalis KI-67c
functionalis mitotic indexd
basalis mitotic index
basalis KI-67c
1760 ( 517
105 ( 34.0
2.2 ( 1.0
0
0
0
5.5 ( 1.15
39 ( 5
a Luteal-phase animals represent values from animals used under other
studies.34,35 Unless otherwise indicated values represent mean ( SE.
b Endometrial and myometrial weight indicate the weight of 1/2 of the uterus.
c KI-67 index represents the number of KI-67 labeled cells per 1000 cells
counted. d Mitotic index represents the number of mitotic cells per 1000
cells counted.
5-(2-Oxoindolin-5-yl)-1H-pyrrole-2-carbonitrile (12). Step
1. tert-Butyl 2-(2-oxoindolin-5-yl)-1H-pyrrole-1-carboxylate. A
mixture of 5-bromo-oxindole (1.34 g, 6.35 mmol), 1-(tert-butoxy-
carbonyl)-1H-pyrrol-2-ylboronic acid (2.67 g, 12.7 mmol), and
sodium carbonate (2.70 g, 25.4 mmol) in THF (75 mL) and water
(25 mL) was degassed by passage of nitrogen gas. To this mixture
was added tetrakis(triphenylphoshine)palladium(0) (0.30 g), and the
mixture heated under reflux (16 h). The reaction mixture was cooled
to room temperature, then poured into water and extracted into
EtOAc, dried (MgSO4), and evaporated. The residue was then
purified by silica gel column chromatography (hexanes/acetone 100:
0-50:50) to afford the product (0.89 g, 47%). 1H NMR (400 MHz,
DMSO-d6) δ 10.36 (br s, 1H), 7.24 (dd, J ) 3.3 and 1.8 Hz), 7.09
(s, 1H), 7.05 (dd, J ) 7.9 and 1.8 Hz, 1H), 6.75 (d, J ) 7.9 Hz,
1H), 6.18 (t, J ) 3.3 Hz, 1H), 6.10 (dd, J ) 3.3 and 1.8 Hz), 3.43
(s, 2H), and 1.27 (s, 9H). MS (ESI, [M + H]+) 299.1.
Step 2. tert-Butyl 2-cyano-5-(2-oxoindolin-5-yl)-1H-pyrrole-
1-carboxylate. To a solution of the last cited product (0.85 g, 2.89
mmol) in dry CH2Cl2 (30 mL) at -78 °C was added chlorosulfonyl
isocyanate (0.21 mL, 2.38 mmol) dropwise. After 1 h, dry DMF
(1.3 mL) was added dropwise; the reaction was allowed to warm
up to room temperature. After 1 h, the mixture was poured into
saturated sodium hydrogen carbonate solution, washed with water,
dried (MgSO4), and evaporated. The residue was then purified by
silica gel column chromatography (hexanes/acetone 100:0-0:100)
to afford the product (0.25 g, 26%). 1H NMR (400 MHz, DMSO-
d6) δ 10.46 (br s, 1H), 7.23 (d, J ) 3.7 Hz, 1H), 7.19 (s, 1H),
5-(3,3-Dimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-methyl-
1H-pyrrole-2-carbonitrile (18). Step 1. 3,3-Dimethylindolin-2-
one. Dimethylindolin-2-one was prepared according to a reported
procedure.23 1H NMR (400 MHz, DMSO-d6) δ 10.24 (br s, 1H),
7.21 (d, J ) 7.4 Hz, 1H), 7.10 (dt, J ) 7.7 and 1.3 Hz, 1H), 7.08
(dt, J ) 7.4 and 0.9 Hz, 1H), 6.78 (d, J ) 7.7 Hz, 1H), and 1.18
(s, 6H).
Step 2. 5-Bromo-3,3-dimethylindolin-2-one. 3,3-Dimethyl-
indol-2-one (0.65 g, 4.03 mmol) and sodium acetate (0.33 g, 4.07
mmol) were stirred in acetic acid (5 mL); then bromine (0.66 g,
4.13 mmol) in acetic acid (5 cm3) was added dropwise to the
reaction mixture. The reaction was stirred for 50 min and then
poured into water. The mixture was basified with sodium carbonate
and then extracted with ethyl acetate (×3), dried (MgSO4), filtered,
and evaporated to provide the title compound, which required no
1
further purification (0.89 g, 92%). H NMR (400 MHz, DMSO-
d6) δ 10.38 (br s, 1H), 7.47 (d, J ) 2.0 Hz, 1H), 7.28 (dd, J ) 8.2
and 2.0 Hz, 1H), 6.75 (d, J ) 8.2 Hz, 1H), and 1.20 (s, 6H).
Step 3. 5-(3,3-Dimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-
methyl-1H-pyrrole-2-carbonitrile (18). The title compound was
prepared from 5-bromo-3,3-dimethylindolin-2-one (14.4 g, 60
mmol) according to the procedure described for compound (13)
and purified by silica gel column chromatography (hexanes/EtOAc
3:1-1:1) to afford the title compound (18) (1.76 g, 11%). 1H NMR

1868 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Fensome et al.
(400 MHz, DMSO-d6) δ 10.50 (s, 1H), 7.47 (s, 1H), 7.30 (dd, J )
8.0 and 1.9 Hz, 1H), 7.02 (d, J ) 3.8 Hz, 1H), 6.95 (d, J ) 8.0 Hz,
1H), 6.35 (d, J ) 3.7 Hz, 1H), 3.71 (s, 3H), and 1.30 (s, 6H). HRMS
calculated for C16H15N3O + H+ 266.12934; found (ESI, [M + H]+)
266.1298.
3H), 1.98-1.93 (m, 2H), 1.86–1.77 (m, 2H), and 0.69 (t, J ) 7.3
Hz, 6H). HRMS calculated for C18H19N3O + H+ 294.16009; found
(ESI, [M + H]+) 294.1616.
1-Methyl-5-(2′-oxo-1′,2′-dihydrospiro[cyclopentane-1,3′-indol]-
5′-yl)-1H-pyrrole-2-carbonitrile (20). Step 1. Spiro[cyclopen-
tane-1,3′-[3H]indol]-2′(1′H)-one. Spiro[cyclopentane-1,3′-[3H]indol]-
2′(1′H)-one was prepared according to a described procedure23 from
oxindole (2.0 g, 15.0 mmol) to afford the product (1.4 g, 50%) as
a tan solid. 1H NMR (300 MHz, CDCl3) δ 9.30 (br s, 1H),
7.25-7.14 (m, 2H), 7.01 (dd, J ) 7.5 and 1.0 Hz, 1H), 6.94 (dd,
J ) 7.5 and 1.0 Hz, 1H), and 2.2-1.80 (m, 8H).
5-(3,3-Diethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-methyl-1H-
pyrrole-2-carbonitrile (19). Step 1. 3-Ethyl-indol-2-one. A solu-
tion of oxindole (40 g, 0.3 mol) in dry THF (400 mL) under N2
was cooled to -25 °C and treated dropwise with n-butyl lithium
(2.5 M in hexanes, 240 mL, 0.6 mol). To the resulting solution
was added N,N,N′,N′-tetramethylethylenediamine (90.4 mL, 0.6
mol). After 30 min, iodoethane (48 mL, 0.6 mol) was added, and
the reaction mixture was allowed to warm to room temperature.
After 16 h, the reaction was quenched (saturated NH4Cl solution),
extracted with EtOAc, washed with diluted HCl, water, and brine,
dried (MgSO4), and evaporated. The residual oil was triturated with
hexane to afford the crude product (24.5 g, 51%). A sample (3 g)
was recrystallized from EtOAc/hexane to obtain the title compound
Step 2. 5-Bromo-spiro[cyclopentane-1,3′-[3H]indol]-2′(1′H)-
one. Prepared following the procedure for compound 18, step 2,
from spiro[cyclopentane-1,3′-[3H]indol]-2′(1′H)-one (0.27 g, 1.4
mmol) to afford the title compound (0.37 g, 96%) as an off-white
1
solid, which was used without further purification. H NMR (300
MHz, CDCl3) δ 8.63 (br s, 1H), 7.39–7.30 (m, 2H), 6.79 (d, J )
8.0 Hz, 1H), and 2.27-1.80 (m, 8H).
1
Step 3. 1-Methyl-5-(2′-oxo-1′,2′-dihydrospiro[cyclopentane-
1,3′-indol]-5′-yl)-1H-pyrrole-2-carbonitrile (20). Compound 20
was prepared according the procedure for compound 19, step 5,
from 5-bromo-spiro[cyclopentane-1,3′-[3H]indol]-2′(1′H)-one (0.30
g, 1.12 mmol) and purified by silica gel column chromatography
(hexanes/acetone 100:0-0:100) to afford the title compound 20
(1.4 g). H NMR (400 MHz, DMSO-d6) δ 10.3 (s, 1H), 7.22 (m,
1H), 7.15 (m, 1H), 6.93 (dt, J ) 7.45 and 1.10 Hz, 1H), 6.8 (dt, J
) 7.69 and 0.45 Hz, 1H), 3.38 (t, J ) 5.7 Hz, 3H), 1.8–2.0 (m,
2H), and 0.76 (t, J ) 7.5 Hz, 3H). MS (ESI, [M + H]) m/z 270.
Step 2. 3,3-Diethyl-indol-2-one. 3-Ethylindol-2-one (16 g, 0.1
mol) was resubjected to the conditions described in the preceding
experiment to afford the title product (9.0 g, 45%), mp 156–159
°C. 1H NMR (400 MHz, DMSO-d6) δ 10.44 (s,1H), 7.70-7.69 (t,
1H), 7.62-7.59 (m, 1H), 7.58 (d,1H J ) 1.7 Hz), 7.53–7.50 (m,
1H), 7.45-7.41 (t,1H), 7.36-7.35 (m, 1H), 7.34-7.33 (m, 1H),
6.91-6.89 (d, 1H J ) 8.2 Hz), 1.87-1.80 (m, 2H), 1.77-1.70 (m,
1
(0.23 g, 72%). H NMR (400 MHz, DMSO-d6) δ 10.44 (s, 1H),
7.36 (d, J ) 1.6 Hz, 1H), 7.28 (dd, J ) 7.9 and 1.7 Hz, 1H), 7.00
(d, J ) 4.0 Hz, 1H), 6.92 (d, J ) 8.0 Hz, 1H), 6.29 (d, J ) 4.0 Hz,
1H), 3.70 (s, 3H), 2.02-1.93 (m, 6H), and 1.84-1.80 (m, 2H).
HRMS calculated for C18H17N3O + H+ 292.1444; found (ESI, [M
+ H]+) 292.1463.
1
2H), and 0.54-0.50 (t, 6H). MS (+ESI, [M + H]) m/z 190. H
NMR (400 MHz, DMSO-d6) δ 10.32 (br s, 1H), 7.18-7.14 (m,
2H), 6.99 (dt, J ) 10.7 and 1.0 Hz, 1H), 6.83 (dd, J ) 7.9 and 0.8
Hz, 1H), 1.76–1.67 (m, 4H), and 0.50 (t, J ) 7.4 Hz, 6H). MS
(+ESI, [M + H]+) m/z 190.
5-(Spiro[cyclohexane-1,3′-[3H]indole]-2′-oxo-5′-yl)-1H-pyrrole-
1-methyl-2-carbonitrile (21). Step 1. Spiro[cyclohexane-1,3′-
[3H]indol]-2′-(1′H)one. Spiro[cyclohexane-1,3′-[3H]indol]-2′-
(1′H)one was prepared according to a described procedure23 from
oxindole (25.0 g, 0.19 mol) to afford the product (26.3 g, 69.6%)
as colorless crystals, mp 110–114 °C. 1H NMR (300 MHz, DMSO-
d6) δ 10.3 (br s, 1H), 7.44 (d, J ) 8.0 Hz, 1H), 7.17 (t, J ) 8 Hz,
7.17 (t, 1H, J ) 8 Hz), 6.94 (t, J ) 8.0 Hz, 1H), 6.84 (d, 1H, J )
8.0 Hz, 1H), and 1.67 (m, 10H).
Step 3. 5-Bromo-3,3-diethyl-1,3-dihydro-indol-2-one. 5-Bromo-
3,3-diethyl-1,3-dihydro-indol-2-one was prepared following the
procedure for compound 18 step 2, from 3,3-diethyl-1,3-dihydro-
indol-2-one (0.5 g, 2.6 mmol) affording the product, which was
used without further purification (0.6 g, 85%), as a light yellow
1
solid. H NMR (400 MHz, DMSO-δ6) d 10.45 (s, 1H), 7.40 (d, J
Step 2. 5′-Bromospiro[cyclohexane-1,3′-[3H] indol]-2′(1′H)-
one. 5′-Bromospiro[cyclohexane-1,3′-[3H] indol]-2′(1′H)-one was
prepared following the procedure for compound 18, step 2, from
spiro[cyclohexane-1,3′-[3H]indol]-2′(1′H)-one (17.6 g, 0.09 mol)
to afford the product (16.5 g, 67%) as off-white crystals, mp 196
) 2.2 Hz, 1H), 7.34-7.31 (m, 1H), 6.77 (d, 1H J ) 8.2 Hz, 1H),
1.78–1.65 (m, 4H), and 0.50-0.46 (m, 6H). MS (-ESI [M - H])
m/z 266/268.
Step 4. 5-Cyano-1-methyl-1H-pyrrol-2-ylboronic acid (16). To
a solution of 1-methyl-1H-pyrrole-2-carbonitrile (1.20 g, 11.3 mmol)
in dry THF (35 mL) at 0 °C was added tri-iso-propyl borate (2.6
mL, 11.3 mmol) and lithium di-iso-propylamine (7.3 mL, 14.7
mmol 2.0 M, in THF/hexane/ethylbenzene). The reaction was
allowed to warm to room temperature and stirred (2 h). The mixture
was quenched (saturated aq NH4Cl) and extracted with EtOAc (3×).
The organic layer was dried (Na2SO4), and evaporated (<25 °C)
to give the product, which was used without further purification.
1H NMR (300 MHz, DMSO-d6) δ 8.37 (bs, 2H), 6.87 (d, J ) 4.0
Hz, 1H), 6.77 (d, J ) 4.1 Hz, 1H), and 3.88 (s, 3H).
Step 5. 5-(3,3-Diethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-
methyl-1H-pyrrole-2-carbonitrile (19). To a solution of 5-bromo-
3,3-diethyl-1,3-dihydro-indol-2-one (0.6 g, 0.20 mmol) in dry THF
(55 mL) was added tetrakis(triphenylphosphine)palladium(0) (0.26
g) after evacuation and purging with nitrogen. After 20 min,
potassium carbonate (1.5 g, 11.1 mmol) and the above-prepared
boronic acid were added, followed by water (13 mL). The mixture
was heated under reflux (16 h). The reaction mixture was cooled
and filtered through Celite, which was washed with EtOAc. The
combined organic layers were washed with water and brine, then
dried (Na2SO4) and evaporated. The residue was then purified by
silica gel column chromatography (MeOH/CH2Cl2, 1:99–4:99)
followed by further purification by silica gel column chromatog-
raphy (hexanes/EtOAc 5:1-2:1) to give the title compound (19)
(320 mg, 49%). 1H NMR (400 MHz, CDCl3) δ 7.74 (bs, 1H), 7.23
(dd, J ) 7.9 and 1.8 Hz, 1H), 7.12 (s, 1H), 6.96 (d, J ) 7.9 Hz,
1H), 6.85 (d, J ) 4.0 Hz, 1H), 6.21 (d, J ) 4.0 Hz, 2H), 3.72 (s,
1
-199 °C. H NMR (400 MHz, DMSO-d6) δ 1.62 (m, 10H), 6.8
(d, 1H, J ) 6.8 Hz), 7.36 (d, 1H, J ) 8.2, 1.8 Hz), 7.58 (dd, 1H,
J ) 8.2, 1.8 Hz), 10.44 (S, 1H), 10.44 (s, 1H), 7.58 (dd, J ) 8.2
and 1.8 Hz, 1H), 7.36 (d, J ) 8.2 and 1.8 Hz, 1H), 6.80 (d, J )
6.8 Hz, 1H), and 1.62 (m, 10H).
Step 3. 5-(Spiro[cyclohexane-1,3′-[3H]indole]-2′-oxo-5′-yl)-
1H-pyrrole-1-methyl-2-carbonitrile (21). The title compound was
prepared from 5′-bromospiro[cyclohexane-1,3′-indolin]-2′-one (0.51
g, 1.8 mmol) according to the procedure for compound 12 and
purified by silica gel column chromatography (hexanes/acetone 100:
0-0:100) to afford the product (21) (0.38 g, 68%) as a white solid.
1H NMR (400 MHz, DMSO-d6) δ 10.47 (s, 1H), 7.55 (d, J ) 1.7
Hz, 1H), 7.31 (dd, J ) 8.0 and 1.8 Hz), 7.01 (d, J ) 4.0 Hz, 1H),
6.94 (d, J ) 8.0 Hz, 1H), 6.29 (d, J ) 4.0 Hz), 3.71 (s, 3H),
1.88-1.85 (m, 2H), and 1.72-1.57 (m, 8H). MS (ESI, [M + H]+)
m/z 304.
5-(4-Fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-
methyl-1H-pyrrole-2-carbonitrile (25). Step 1. 2-(2-Fluoro-6-
nitro-phenyl)-malonic Acid Dimethyl Ester. To a solution of 2,3-
difluoronitrobenzene (9.0 g, 56 mmol) in DMF was added potassium
carbonate (13.8 g, 100 mmol) and dimethylmalonate (6.88 mL, 60
mmol), and the reaction was heated to 65 °C (24 h.). After cooling
to room temperature, the mixture was neutralized (dilute HCl) and
extracted with Et2O; the organic layers were dried (MgSO4) and
evaporated. The crude product was crystallized from hexanes/EtOAc
1
(95:5) to afford the product (6.60 g, 43%). H NMR (400 MHz,
DMSO-d6) δ 7.96-7.94 (m, 1H), 7.78–7.71 (m, 1H), 5.52 (s, 1H),

Pyrrole-Oxindole Progesterone Receptor Modulators
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1869
and 3.70 (s, 6H). HRMS calculated for C11H10FNO6 + H+
272.05649; found (ESI, [M + H]+) 272.0576.
5-(6-Fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-
methyl-1H-pyrrole-2-carbonitrile (26). Step 1. Dimethyl 2-(4-
Fluoro-2-nitrophenyl)malonate. Dimethyl 2-(4-fluoro-2-nitrophe-
nyl)malonate was prepared according to the procedure for compound
25, step 1, from 2,5-difluoronitrobenzene (7.95 g, 70 mmol). The
product was recrystallized from hexanes/CH2Cl2 to afford the
product (7.10 g, 37%) as a solid. 1H NMR (400 MHz, DMSO-d6)
δ 8.06 (dd, J ) 8.7 and 2.7 Hz, 1H), 7.70 (dt, J ) 7.9 and 2.9 Hz,
1H), 7.62 (dd, J ) 8.7 and 5.6 Hz, 1H), 5.50 (s, 1H), and 3.70 (s,
6H). HRMS calculated for C11H10FNO6 + H+ 272.05649; found
(ESI, [M + H]+) 272.0561.
Step 2. (2-Fluoro-6-nitrophenyl)acetic acid. 2-(2-Fluoro-6-
nitro-phenyl)-malonic acid dimethyl ester (6.50 g, 23.98 mmol) and
hydrochloric acid (200 mL, 6 N) were heated under reflux (24 h).
After the mixture was cooled to room temperature, the solid was
collected, washed with water, and dried in Vacuo to give the product
1
(3.30 g, 54%). H NMR (400 MHz, DMSO-d6) 12.76 (br s, 1H),
7.95 (dd, J ) 8.1 and 1.2 Hz, 1H), 7.72-7.59 (m 2H), and 3.91 (s,
2H). HRMS calculated for C11H10FNO6 + H+ 272.05649; found
(ESI, [M + H]+) 272.0576.
Step 2. 2-(4-Fluoro-2-nitrophenyl)acetic acid. 2-(4-Fluoro-2-
nitrophenyl)acetic acid was prepared according to the procedure
for compound 25, step 2, from dimethyl 2-(4-fluoro-2-nitrophe-
nyl)malonate (7.10 g, 26.1 mmol). The isolated product (3.94 g,
Step 3. 4-Fluoro-1,3-dihydro-2H-indol-2-one. 2-Fluoro-6-ni-
trophenyl)acetic acid (3.30 g, 16.6 mmol) was dissolved in acetic
acid (20 mL) and hydrogenated (50 psi) over palladium on carbon
(10%, 0.5 g). After 24 h, the catalyst was removed by filtation
through Celite, the filter pad was washed with methanol, and the
combined organics were evaporated. The mixture was then dis-
solved in ethanol (100 mL), treated with para-toluenesulfonic acid
(0.05 g), and heated under reflux (1 h). The cooled mixture was
poured into water, extracted with EtOAc, dried (MgSO4), and
evaporated. The solid was triturated (hexanes/EtOAc 95:5) to afford
the product (1.70 g, 67%). 1H NMR (400 MHz, DMSO-d6) δ 10.53
(s, 1H, NH), 7.19–7.13 (m, 1H), 6.72 (t, J ) 8.59, 1H), 6.61 (d, J
) 7.81, 1H), and 3.48 (s, 2H). HRMS calculated for C8H6FNO +
H+ 152.05062; found (ESI, [M + H]+) 152.0525.
1
75%) was used without further purification. H NMR (400 MHz,
DMSO-d6) δ 12.58 (s, 1H), 7.99 (dt, J ) 8.0 and 2.2 Hz, 1H),
7.67-7.60 (m, 2H), and 3.99 (s, 2H).
Step 3. 6-Fluoroindolin-2-one. 6-Fluoroindolin-2-one was pre-
pared according to the procedure for compound 25, step 3, from
2-(4-fluoro-2-nitrophenyl)acetic acid (3.94 g, 19.79 mmol). The
isolated product (2.67 g, 89%) was used without further purification.
1H NMR (400 MHz, DMSO-d6) δ 10.49 (s, 1H), 7.22-7.18 (m,
1H), 6.72 (dt, J ) 9.1 and 2.5 Hz, 1H), 6.61 (dd, J ) 9.4 and 2.5
Hz), and 3.44 (s, 2H). HRMS calculated for C8H6FNO + H+
152.05062; found (ESI, [M + H]+) 152.0567.
Step 4. 6-Fluoro-3,3-dimethylindolin-2-one. 6-Fluoro-3,3-di-
methylindolin-2-one was prepared according to the procedure for
compound 25, step 4, from 6-fluoroindolin-2-one (1.52 g, 10 mmol).
The residue was purified by silica gel column chromatography
(hexanes/EtOAc 80:20) to afford the product (0.47 g, 23%). 1H
NMR (400 MHz, DMSO-d6) δ 10.45 (s, 1H), 7.29 (dd, J ) 8.2
and 5.6 Hz, 1H), 6.78-6.72 (m, 1H), 6.65 (dd, J ) 9.3 and 2.5
Hz, 1H), and 1.23 (s, 6H).
Step 5. 5-Bromo-6-fluoro-3,3-dimethylindolin-2-one. 5-Bromo-
6-fluoro-3,3-dimethylindolin-2-one was prepared according to the
procedure for compound 25, step 5, from 6-fluoro-3,3-dimethylin-
dolin-2-one (0.67 g, 3.76 mmol). The isolated product (0.88 g, 90%)
was used without further purification. 1H NMR (400 MHz, DMSO-
d6) δ 10.58 (s, 1H), 7.66 (d, J ) 6.9 Hz, 1H), 6.82 (d, J ) 9.0 Hz,
1H), and 1.25 (s, 6H). HRMS calculated for C10H10FNO 179.07464;
found (ESI, M+) 179.0742.
Step 4. 4-Fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-2-one.
4-Fluoro-1,3-dihydro-2H-indol-2-one (3.40 g, 22.5 mmol) and
lithium chloride (2.7 g, 60 mmol) were dissolved in THF (100 mL).
The solution was cooled (-78 °C), and n-butyl lithium (7 mL, 2.5
M in hexanes, 15 mmol) was added slowly over 15 min. Methyl
iodide (3.1 mL, 50 mmol) was added, and the mixture allowed to
warm up to room temperature. After 16 h, the mixture was poured
into water and extracted with EtOAc; the organic layer was dried
(MgSO4) and evaporated. The resiude was purified by silica gel
column chromatography (hexanes/EtOAc 90:10-80:20) to afford
the product (1.00 g, 25%). 1H NMR (400 MHz, DMSO-d6) δ 10.53
(s, 1H), 7.20–7.15 (m, 1H), 6.72 (t, J ) 8.84, 1H), 6.67 (d, J )
7.66, 1H), and 1.29 (s, 6H). HRMS calculated for C10H10FNO +
H+ 180.08192; found (ESI, [M + H]+) 180.0816.
Step 5. 5-Bromo-4-fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-
2-one. 4-Fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-2-one (1.00 g,
22.9 mmol) was dissolved in CH2Cl2 (50 mL) and acetic acid (2
mL) was added. Bromine (0.38 mL, 7.5 mmol) was added dropwise,
and the solution was allowed to stir (16 h). The reaction mixture
was poured into saturated sodium thiosulfate solution, extracted
with Et2O, dried (MgSO4), and evaporated. The residue was washed
Step 6. 5-(6-Fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-
5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (26). Compound 26 was
prepared following the procedure for compound 12 from 5-bromo-
6-fluoro-3,3-dimethylindolin-2-one (0.88 g, 3.40 mmol). The residue
was purified by silica gel column chromatography (hexanes/EtOAc
1
80:20-75:25) to afford the title compound 26 (0.27 g, 28%). H
1
NMR (400 MHz, DMSO-d6) δ 10.66 (s, 1H), 7.41 (d, J ) 7.4 Hz,
1H), 7.04 (d, J ) 4.0 Hz, 1H), 6.81 (d, J ) 10.1 Hz, 1H), 6.29 (d,
J ) 4.0 Hz, 1H), 3.59 (d, J ) 1.2 Hz, 3H), and 1.28 (s, 6H). HRMS
calculated for C16H14FN3O + H+ 284.11937; found (ESI, [M +
H]+) 284.1206.
with hexane to afford the product (1.25 g, 87%). H NMR (400
MHz, DMSO-d6) δ 10.70 (s, 1H), 7.5 (dd, J ) 7.15 and 1.04 Hz,
1H), 6.69 (d,J ) 8.31 Hz, 1H), and 1.34 (s, 6H). HRMS calculated
for C10H9BrFNO + H+ 257.99243; found (ESI, [M + H]+)
257.9936.
5-(7-Fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-
methyl-1H-pyrrole-2-carbonitrile (27). Step 1. 2-(3-Fluoro-2-
nitro-phenyl)-malonic acid dimethyl ester. 2-(3-Fluoro-2-nitro-
phenyl)-malonic acid dimethyl ester was prepared following the
procedure for compound 25, step 1, from 2,6-difluoronitrobenzene
(5.00 g, 31.44 mmol). Crystallization from hexanes/EtOAc (95:5)
Step 6. 5-(4-Fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-
5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (25). 5-Bromo-4-fluoro-
3,3-dimethyl-1,3-dihydro-2H-indol-2-one (1.25 g, 4.86 mmol) and
tetrakis(triphenylphosphine)palladium(0) (0.4 g) were dissolved in
ethylene glycol dimethyl ether (40 mL) and stirred 15 min.
N-Methyl-5-cyanopyrroleboronic acid (2.00 g, 13.33 mmol) and
potassium carbonate (3.48 g, 25 mmol) were added, followed by
water (20 mL), and the mixture was heated under reflux (16 h).
After cooling to room temperature, the mixture was poured into
water, neutralized (dilute HCl), and extracted with EtOAc. The
organic layer was dried (MgSO4) and evaporated. The product was
purified by silica gel column chromatography (hexanes/THF 90:
10-70:30) to afford the title compound (25) (0.060 g, 5%). 1H
NMR (400 MHz, DMSO-d6) δ 10.72 (s, 1H), 7.25 (t, J ) 7.8,
1H), 7.01 (d, J ) 4.1, 1H), 6.79 (d, J ) 7.9 Hz, 1H), 6.26 (d, J )
4.00 Hz, 1H), 3.56 (s, 3H), and 1.34 (s, 6H). HRMS calcd for
C16H14FN3O + H+ 284.11937; found (ESI, [M + H]+) 284.1193.
1
afforded the product (4.60 g, 54%). H NMR (400 MHz, DMSO-
d6) δ 7.81–7.75 (m, 1H), 7.69–7.64 (m, 1H,), 7.43 (d, J ) 7.93,
1H), 5.32 (s, 1H), and 3.71 (s, 6H). HRMS calculated for
C11H10FNO6 271.0492; found (ESI, [M + H]+) 272.0576.
Step 2. (3-Fluoro-2-nitro-phenyl)-acetic acid. (3-Fluoro-2-nitro-
phenyl)-acetic acid was prepared following the procedure for
compound 25, step 2, from 2-(3-fluoro-2-nitro-phenyl)-malonic acid
dimethyl ester (12.00 g, 44 mmol). Crystallization from hexanes/
EtOAc (95:5) gave the product (7.60 g, 54%). 1H NMR (400 MHz,
DMSO-d6) δ 12.69 (s, 1H), 7.62–7.67 (m, 1H), 7.47–7.52 (m, 1H),
7.35 (d, J ) 7.80, 1H), and 3.84 (s, 2H). MS (ESI, [M - H]-) m/z
197.9.

1870 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Fensome et al.
Step 3. 7-Fluoro-1,3-dihydro-indol-2-one. 7-Fluoro-1,3-dihy-
dro-indol-2-one was prepared following the procedure for com-
pound 25, step 3, from (3-fluoro-2-nitro-phenyl)-acetic acid (9.60
g, 48 mmol). The residue was triturated with hexanes/EtOAc (95:
5) to afford the product (6.00 g, 83%). 1H NMR (400 MHz, DMSO-
d6) δ 10.83 (s, 1H, NH), 7.05–7.10 (m, 2H, ArH), 6.91–6.96 (m,
1H, ArH), and 3.56 (s, 2H). HRMS calculated for C8H6FNO +
H+ 151.0433; found (ESI, [M + H]+) 152.0515.
(s, 1H, NH), 7.44 (dd, J ) 9.75 Hz and1.69 Hz, 1H), 7.36 (d, J )
1.7 Hz, 1H), 1.91-1.68 (m, 4H), and 0.51 (t, J ) 7.4 Hz, 6H).
HRMS calculated for C12H13BrFNO + H+ 286.02373; found (ESI,
[M + H]+) 286.024.
Step 3. 5-(7-Fluoro-3,3-diethyl-2-oxo-2,3-dihydro-1H-indol-
5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (28). In a 40 mL septa
cap vial, 5-bromo-7-fluoro-3,3-diethyl-1,3-dihydro-2H-indol-2-one
(0.48 g, 1.67 mmol), tris(dibenzylideneacetone) dipalladium(0)
chloroform (0.042 g, 0.042 mmol), N-methyl-5-cyanopyrrole-2-
boronic acid (0.54 g, 3.6 mmol), and potassium fluoride (0.83 g,
6.6 mmol) were combined, and THF (5 mL) was added under a
nitrogen atmosphere. Tri-tert-butyl phosphine (0.24 mL, 0.082
mmol, 10% in hexane) was added. After 3 h, hexanes/EtOAc (50:
50) was added, and the reaction mixture was filtered through a pad
of silica gel. The solvent was evaporated, and the solid was
recrystallized from hexanes/EtOAc to give the title compound 28
Step 4. 7-Fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-2-one.
7-Fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-2-one was prepared
following the procedure for compound 25, step 4, from 7-fluoro-
1,3-dihydro-indol-2-one (7.30 g, 48 mmol). The residue was purified
by silica gel column chromatography (hexanes/EtOAc 90:10-80:
20) to afford the product (4.10 g, 48%). 1H NMR (400 MHz,
DMSO-d6) δ 10.81 (s, 1H), 7.15 (d, J ) 7.1, 1H), 7.05-7.10 (m,
1H), 6.95-7.00 (m, 1H), and 1.26 (s, 6H). HRMS calculated for
C10H10FNO + H+ 180.08192; found (ESI, [M + H]+) 180.0831.
Step 5. 5-Bromo-7-fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-
2-one. 5-Bromo-7-fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-2-one
was prepared following the procedure for compound 25, step 5,
from 7-fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-2-one (4.1 g, 22.9
mmol). The residue was triturated with hexanes to afford the product
1
(0.25 g, 48%). H NMR (400 MHz, DMSO-d6) δ 11.07 (s, 1H),
7.32-7.29 (dd, J ) 11.0 J ) 1.5 Hz, 1H), 7.27 (d, J ) 1.4 Hz,
1H), 7.02 (d, J ) 4.1 Hz, 1H), 6.37 (d, J ) 4.0 Hz, 1H), 3.72 (s,
3H), 1.68–1.87 (m, 4H), and 0.56 (t, J ) 7.4 Hz, 6H). HRMS
calculated for C18H18FN3O + H+ 312.15067; found (ESI, [M +
H]+) 312.1507.
1
(4.84 g, 82%). H NMR (400 MHz, DMSO-d6) δ 10.97 (s, 1H),
5-(7′-Fluoro-2′-oxo-1′,2′-dihydrospiro[cyclopentane-1,3′-indol]-
5′-yl)-1-methyl-1H-pyrrole-2-carbonitrile (29). Step 1. 7′-Fluo-
rospiro[cyclopentane-1,3′-indolin]-2′-one. 7′-Fluorospiro[cyclo-
pentane-1,3′-indolin]-2′-one was prepared following the procedure
for compound 25, step 4, from 7-fluoro-1,3-dihydro-indol-2-one
(1.00 g, 6.61 mmol). The residue was purified by silica gel column
chromatography (hexanes/EtOAc 100:0-0:100) to afford the
7.45 (d, J ) 1.6 Hz, 1H), 7.41 (dd, J ) 9.7 and 1.6 Hz, 1H), and
1.27 (s, 6H). HRMS calculated for C10H9BrFNO - H+ 255.97788;
found (ESI, [M - H]-) 255.9781.
Step 6. 5-(7-Fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-
5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (27). Compound 27 was
prepared following the procedure for compound 19 from 5-bromo-
7-fluoro-3,3-dimethyl-1,3-dihydro-2H-indol-2-one (3.60 g, 14 mmol)
replacing THF with DMF (140 mL) as the solvent. The residue
was purified by silica gel column chromatography (CH2Cl2/EtOAc,
98:2) to afford the title compound 27 (0.75 g, 19%). 1H NMR (400
MHz, DMSO-d6) δ 11.01 (s, 1H), 7.36 (d, J ) 1.4 Hz, 1H), 7.29
(dd, J ) 11.2 and 1.4 Hz, 1H), 7.02 (d, J ) 4.0 Hz, 1H), 6.35 (d,
J ) 4.0 Hz, 1H), 3.73 (s, 3H), and 1.31 (s, 6H). HRMS calculated
for C16H14FN3O - H+ 282.10481; found (ESI, [M - H]-)
282.1034.
1
product (0.59 g, 44%). H NMR (400 MHz, DMSO-d6) δ 10.71
(br s, 1H), 7.04-6.98 (m, 2H), 6.93-6.88 (m, 1H), 1.96-1.84 (m,
6H), and 1.74-1.67 (m, 2H). MS (ESI, [M + H]+) 206.1.
Step 2. 5′-Bromo-7′-fluorospiro[cyclopentane-1,3′-indolin]-
2′-one. 5′-Bromo-7′-fluorospiro[cyclopentane-1,3′-indolin]-2′-one
was prepared following the procedure for compound 25, step 5,
from 7′-fluorospiro[cyclopentane-1,3′-indolin]-2′-one (0.57 g, 2.77
mmol). The product (0.76 g, 91%) was used without further
purification or characterization.
Alternative Preparation of 5-(7-Fluoro-3,3-dimethyl-2-oxo-
2,3-dihydro-1H-indol-5-yl)-1-methyl-1H-pyrrole-2-carboni-
trile (27). 5-Bromo-7-fluoro-3,3-dimethyl-1,3-dihydro-indol-2-one
(5.16 g, 20.0 mmol), 1-methyl-5-cyano-2-pyrroleboronic acid (5.4
g, 36 mmol), KF (3.83 g, 66 mmol), and Pd2(dba)3 monochloroform
adduct (516 mg, 0.500 mmol) were added to a 200 mL round-
bottom flask under nitrogen. The flask was sealed and purged with
nitrogen for 5 min. THF (50 mL) was added, and the mixture was
purged with nitrogen for an additional 5 min. A solution of tri-
tert-butylphosphine (10 wt % in hexanes; 2.97 mL, 1.00 mmol)
was added via syringe, and the mixture was stirred vigorously at
25 °C for 5 h. The mixture was diluted with 250 mL of EtOAc,
filtered through a plug of silica gel, washed through with 200 mL
of EtOAc, and concentrated to give a crude brown/black semisolid.
Purification by flash chromatography (20% acetone/hexane) af-
forded the title compound 27 (4.5 g, 80%) as an off-white solid.
5-(7-Fluoro-3,3-diethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-
methyl-1H-pyrrole-2-carbonitrile (28). Step 1. 7-Fluoro-3,3-
diethyl-1,3-dihydro-2H-indol-2-one. 7-Fluoro-3,3-diethyl-1,3-
dihydro-2H-indol-2-one was prepared following the procedure for
compound 17, step 4, from7-fluoro-1,3-dihydro-indol-2-one (1.00
g, 6.60 mmol). The residue was purified by silica gel column
chromatography (hexanes/EtOAc 90:10-80:20) to afford the
Step 3. 5-(7′-Fluoro-2′-oxo-1′,2′-dihydrospiro[cyclopentane-
1,3′-indol]-5′-yl)-1-methyl-1H-pyrrole-2-carbonitrile (29). Com-
pound 29 was prepared according to the procedure for compound
28, step 3, from 5′-bromo-7′-fluorospiro[cyclopentane-1,3′-indolin]-
2′-one (0.56 g, 1.96 mmol). The residue was purified by silica gel
column chromatography (hexanes/EtOAc 80:20-0:100) to afford
the title compound 29 (0.37 g, 61%). 1H NMR (400 MHz, DMSO-
d6) δ 10.98 (s, 1H), 7.42 (d, J ) 1.3 Hz, 1H), 7.30 (dd, J ) 11.0
and 1.4 Hz, 1H), 7.02 (d, J ) 4.0 Hz, 1H), 6.35 (d, J ) 4.0 Hz,
1H), 3.73 (s, 3H), 1.88-1.86 (m, 2H), and 1.71-1.56 (m, 6H).
HRMS calculated for C18H16FN3O + H+ 310.13502; found (ESI,
[M + H]+) 310.1363.
5-(7′-Fluoro-2′-oxo-1′,2′-dihydrospiro[cyclohexane-1,3′-indol]-
5′-yl)-1-methyl-1H-pyrrole-2-carbonitrile (30). Step 1. 7′-Fluo-
rospiro[cyclohexane-1,3′-indolin]-2′-one. 7′-Fluorospiro[cyclo-
hexane-1,3′-indolin]-2′-one was prepared according to the procedure
for compound 18, step 2, from 7-fluoroindolin-2-one (1.00 g, 6.61
mmol). The residue was purified by silica gel column chromatog-
raphy (hexanes/EtOAc 90:10) and then recrystallized from hexanes
to afford the product (0.40 g, 40%). 1H NMR (400 MHz, DMSO-
d6) δ 10.78 (s, 1H), 7.30 (dd, J ) 7.4 and 0.6 Hz, 1H), 7.12-7.07
(m, 1H), 6.99-6.94 (m, 1H), 1.89-1.82 (m, 2H), 1.89-1.62 (m,
5H), and 1.55-1.49 (m, 3H). HRMS calculated for C13H14FNO +
H+ 220.11322; found (ESI, [M + H]+) 220.1159.
1
product (0.40 g, 29%). H NMR (400 MHz, DMSO) δ 10.86 (s,
1H), 7.14-6.98 (m, 3H), 1.79-1.69 (m, 4H), and 0.51 (t, J ) 7.5
Hz, 6H). HRMS calculated for C12H14FNO + H+ 207.2477; found
(ESI, [M + H]+) 208.1134.
Step 2. 5-Bromo-7-fluoro-3,3-diethyl-1,3-dihydro-2H-indol-
2-one. 5-Bromo-7-fluoro-3,3-diethyl-1,3-dihydro-2H-indol-2-one
was prepared following the procedure for compound 25, step 5,
from 7-fluoro-3,3-diethyl-1,3-dihydro-2H-indol-2-one (0.38 g, 1.83
mmol). The crude residue was triturated with hexanes to afford
the product (0.49 g, 93%). 1H NMR (400 MHz, DMSO-d6) δ 11.03
Step 2. 5′-Bromo-7′-fluorospiro[cyclohexane-1,3′-indolin]-2′-
one. 5′-Bromo-7′-fluorospiro[cyclohexane-1,3′-indolin]-2′-one was
prepared following the procedure for compound 11, step 2, from
7′-fluorospiro[cyclohexane-1,3′-indolin]-2′-one (0.34 g, 1.57 mmol)
to afford the product (0.39 g, 83%), which was used without further
purification or characterization.
Step 3. 5-(7′-Fluoro-2′-oxo-1′,2′-dihydrospiro[cyclohexane-
1,3′-indol]-5′-yl)-1-methyl-1H-pyrrole-2-carbonitrile (30). Com-
pound 30 was prepared according to the procedure for compound

Pyrrole-Oxindole Progesterone Receptor Modulators
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6 1871
28, step 3, from 5′-bromo-7′-fluorospiro[cyclohexane-1,3′-indolin]-
2′-one (0.39 g, 1.30 mmol). The residue was purified by silica gel
column chromatography (hexanes/EtOAc 90:10) to afford the title
HRMS calculated for C21H22FN3O + H+ 352.18197; found (ESI,
[M + H]+) 352.184.
5-(1-Cyclohexyl-7-fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-
indol-5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (43). Compound
43 was prepared following the procedure for compound 37 using
cyclohexyl iodide (57 µL, 0.5 mmol) to afford the title compound
43 (0.004 g, 3%). 1H NMR (400 MHz, DMSO-d6) δ 7.44 (d, J )
1.7 Hz, 1H), 7.36 (dd, J ) 13.4 and 1.6 Hz, 1H), 7.04 (d, J ) 4.1
Hz, 1H), 6.38 (d, J ) 4.0 Hz, 1H), 4.23-4.37 (m, 1H), 3.75 (s,
3H), 2.09-1.98 (m, 2H), 1.85-1.82 (m, 2H), 1.73-1.65 (m, 2H),
1.41-1.32 (m, 8H), and 1.27-1.14. HRMS calculated for
C22H24FN3O + H+ 366.19762; found (ESI, [M + H]+) 366.1978.
1
compound 30 (0.078 g, 18%). H NMR (400 MHz, DMSO-d6) δ
10.96 (s, 1H), 7.28 (dd, J ) 11.0 and 1.6 Hz, 1H), 7.24 (d, J ) 1.4
Hz, 1H), 7.02 (d, J ) 4.0 Hz, 1H), 6.36 (d, J ) 4.0 Hz, 1H), 3.72
(s, 3H), and 2.02-1.84 (m, 8H). HRMS calculated for C19H18FN3O
+ H+ 324.15067; found (ESI, [M + H]+) 324.1517.
5-(7-Fluoro-1,3,3-trimethyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-
1-methyl-1H-pyrrole-2-carbonitrile (37). To 5-(7-fluoro-3,3-di-
methyl-2-oxo-2,3-dihydro-1H-indol-5-yl)-1-methyl-1H-pyrrole-2-
carbonitrile 27 (0.100 g, 0.35 mmol) in dry THF (2 mL) under
nitrogen was added potassium tert-butoxide (0.5 mL, 1 N in THF,
0.5 mmol). After 15 min, methyl iodide (31 µL, 0.5 mmol) was
added, and the mixture stirred (16 h). After evaporation, the residue
was purified by silica gel column chromatography (hexanes/EtOAc
100:0-0:100) to afford the title compound 37 (0.083 g, 79%). 1H
NMR (400 MHz, DMSO-d6) δ 7.42 (d, J ) 1.6 Hz, 1H), 7.36 (dd,
J ) 12.6 and 1.6 Hz, 1H), 7.03 (d, J ) 4.0 Hz, 1H), 6.38 (d, J )
4.1 Hz, 1H), 3.74 (s, 3H), 3.33 (d, J ) 2.7 Hz, 3H), and 1.33 (s,
6H). HRMS calculated for C17H16FN3O + H+ 298.13502; found
(ESI, [M + H]+) 298.1366.
5-(1-Allyl-7-fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-
5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (44). Compound 44 was
prepared following the procedure for compound 37 using allyl
iodide (45 µL, 0.5 mmol) to afford the title compound 44 (0.077
1
g, 68%). H NMR (400 MHz, DMSO-d6) δ 7.46 (d, J ) 1.6 Hz,
1H), 7.34 (dd, J ) 12.5 and 1.6 Hz, 1H), 7.03 (d, J ) 4.1 Hz, 1H),
6.38 (d, J ) 4.0 Hz, 1H), 5.99-5.91 (m, 1H), 5.16 (d, J ) 1.3 Hz,
1H), 5.13 (d, J ) 1.3 Hz, 1H), 4.41 (br d, J ) 3.5 Hz, 2H), 3.74
(s, 3H), and 1.36 (s, 6H). HRMS calculated for C19H18FN3O +
H+ 324.15067; found (ESI, [M + H]+) 324.1512.
5-(1-Ethyl-7-fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-indol-
5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (38). Compound 38 was
prepared following the procedure for compound 37 using ethyl
iodide (40 µL, 0.5 mmol) to afford the title compound 38 (0.072
5-(7-Fluoro-3,3-dimethyl-2-oxo-1-prop-2-yn-1-yl-2,3-dihydro-
1H-indol-5-yl)-1- methyl-1H-pyrrole-2-carbonitrile (45). Com-
pound 45 was prepared following the procedure for compound 37
using propargyl bromide (45 µL, 0.5 mmol) to afford the title
1
1
g, 65%). H NMR (400 MHz, DMSO-d6) δ 7.44 (d, J ) 1.6 Hz,
compound 45 (0.050 g, 43%) H NMR (400 MHz, DMSO-d6) δ
1H), 7.37 (dd, J ) 12.7 and 1.6 Hz, 1H), 7.03 (d, J ) 4.0 Hz, 1H),
6.38 (d, J ) 4.0 Hz, 1H), 3.81 (q, J ) 14.1 and 7.3 Hz, 2H), 3.74
(s, 3H), 1.33 (s, 6H), and 1.21 (t, J ) 6.9 Hz, 3H). HRMS calculated
for C18H18FN3O + H+ 312.15067; found (ESI, [M + H]+)
312.1524.
7.47 (d, J ) 1.6 Hz, 1H), 7.40 (dd, J ) 12.2 and 1.4 Hz, 1H), 7.04
(d, J ) 4.0 Hz, 1H), 6.39 (d, J ) 4.1 Hz, 1H), 4.58 (d, J ) 1.9 Hz,
2H), 3.75 (s, 3H), 3.29 (t, J ) 2.2 Hz), and 1.36 (s, 6H). HRMS
calculated for C19H16FN3O + H+ 322.13502; found (ESI, [M +
H]+) 322.1350.
5-(7-Fluoro-3,3-dimethyl-2-oxo-1-propyl-2,3-dihydro-1H-in-
dol-5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (39). Compound 39
was prepared following the procedure for compound 37 using
1-iodopropane (49 µL, 0.5 mmol) to afford the title compound 39
5-(1-Benzyl-7-fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-in-
dol-5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (46). Compound 46
was prepared following the procedure for compound 37 using
benzyl bromide (59 µL, 0.5 mmol) to afford the title compound 46
1
1
(0.070 g, 58%) H NMR (400 MHz, DMSO-d6) δ 7.44 (d, J )
(0.077 g, 57%). H NMR (400 MHz, DMSO-d6) δ 7.47 (d, J )
1.56 Hz, 1H), 7.36 (dd, J ) 12.7 and 1.6 Hz, 1H), 7.03 (d, J ) 4.0
Hz, 1H), 6.38 (d, J ) 4.0 Hz, 1H), 3.77-3.74 (m, 5H), 1.68-1.62(m,
2H), 1.34 (s, 6H), and 0.87 (t, J ) 7.4 Hz, 3H). HRMS calculated
for C19H20FN3O + H+ 326.16632; found (ESI, [M + H]+)
326.1652.
1.6 Hz, 1H), 7.37-7.33 (m, 2H), 7.30-7.22 (m, 4H), 7.02 (d, J )
4.1 Hz, 1H), 6.36 (d, J ) 4.0 Hz, 1H), 6.36 (s, 2H), 5.01 (s, 3H),
3.72 (s, 3H), and 1.41 (s, 6H). HRMS calculated for C23H20FN3O
+ H+ 374.16632; found (ESI, [M + H]+) 374.1685.
5-[7-Fluoro-3,3-dimethyl-2-oxo-1-(2-phenylethyl)-2,3-dihydro-
1H-indol-5-yl]-1-methyl-1H-pyrrole-2-carbonitrile (47). Com-
pound 47 was prepared following the procedure for compound 37
using phenethyl bromide (67 µL, 0.5 mmol) to afford the title
5-(7-Fluoro-1-isopropyl-3,3-dimethyl-2-oxo-2,3-dihydro-1H-
indol-5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (40). Compound
40 was prepared following the procedure for compound 37 using
2-iodopropane (50 µL, 0.5 mmol) to afford the title compound 40
1
compound 47 (0.041 g, 28%). H NMR (400 MHz, DMSO-d6) δ
1
(0.055 g, 48%). H NMR (400 MHz, DMSO-d6) δ 7.45 (d, J )
7.42 (d, J ) 1.6 Hz, 1H), 7.37 (dd, J ) 12.6 and 1.6 Hz, 1H),
7.28-7.26 (m, 2H), 7.25-7.16 (m, 3H), 7.04 (d, J ) 4.0 Hz), 6.39
(d, J ) 4.1 Hz), 4.01 (t, J ) 7.1 Hz, 2H), 3.75 (s, 3H), 2.93 (t, J
) 7.1 Hz, 2H), and 1.24 (s, 3H). HRMS calculated for C24H22FN3O
+ H+ 388.18197; found (ESI, [M + H]+) 388.1806.
1.6 Hz, 1H), 7.37 (dd, J ) 13.2 and 1.7 Hz, 1H), 7.04 (d, J ) 4.1
Hz, 1H), 6.38 (d, J ) 4.0 Hz, 1H), 4.76-4.73 (m, 1H), 3.75 (s,
3H), 1.39 (d, J ) 7.0 Hz, 6H), and 1.32 (s, 6H). HRMS calculated
for C19H20FN3O + H+ 326.16632; found (ESI, [M + H]+)
326.1661.
Methyl [5-(5-Cyano-1-methyl-1H-pyrrol-2-yl)-7-fluoro-3,3-
dimethyl-2-oxo-2,3-dihydro-1H-indol-1-yl]acetate (48). Com-
pound 48 was prepared following the procedure for compound 37
using methyl bromo acetate (47 µL, 0.5 mmol) to afford the title
5-(7-Fluoro-1-isobutyl-3,3-dimethyl-2-oxo-2,3-dihydro-1H-in-
dol-5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (41). Compound 41
was prepared following the procedure for compound 37 using
2-methyliodopropane (60 µL, 0.5 mmol) to afford the title
1
compound 48 (0.087 g, 68%). H NMR (400 MHz, DMSO-d6) δ
1
compound 41 (0.066 g, 54%). H NMR (400 MHz, DMSO-d6) δ
7.48 (d, J ) 1.5 Hz, 1H), 7.35 (dd, J ) 12.4 and 1.5 Hz, 1H), 7.04
(d, J ) 4.0 Hz), 6.39 (d, J ) 4.0 Hz), 4.66 (s, 1H), 3.75 (s, 3H),
3.72 (s, 3H), and 1.37 (s, 6H). HRMS calculated for C19H18FN3O3
+ H+ 356.14050; found (ESI, [M + H]+) 356.1420.
7.45 (d, J ) 1.6 Hz, 1H), 7.36 (dd, J ) 12.7 and 1.6 Hz, 1H), 7.03
(d, J ) 4.1 Hz, 1H), 6.38 (d, J ) 4.0 Hz, 1H), 3.75 (s, 3H), 3.59
(d, J ) 6.6 Hz, 2H), 2.03-1.99 (m, 1H), 1.34 (s, 6H), and 0.89 (d,
J ) 6.6 Hz, 6H). HRMS calculated for C20H22FN3O + H+
340.18197; found (ESI, [M + H]+) 340.1838.
Biology. T47D Cell Alkaline Phosphatase Assay. T47D Cell
Culture. Frozen T47D cells are thawed in a 37 °C water bath and
diluted to 280 000 cells/mL in culture medium (DMEM/F12 (1:1);
GIBCO, BRL) supplemented with 5% (v/v) charcoal-stripped fetal
bovine serum (not heat-inactivated), 100 U/mL penicillin, 100 µg/
mL streptomycin, and 2 mM GlutaMax (GIBCO, BRL). To each
well in a 96-well plate (Falcon, Becton Dickinson Labware), 180
µL of diluted cell suspension is added. Twenty microliters of
reference or test compounds diluted in the culture medium is then
added to each well. In the antagonist mode, reference antiprogestins
5-(1-Cyclopentyl-7-fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1H-
indol-5-yl)-1-methyl-1H-pyrrole-2-carbonitrile (42). Compound
42 was prepared following the procedure for compound 37 using
cyclopentyliodide (57 µL, 0.5 mmol) to afford the title compound
42 (0.034 g, 27%). 1H NMR (400 MHz, DMSO-d6) δ 7.45 (d, J )
1.7 Hz, 1H), 7.36 (dd, J ) 13.3 and 1.7 Hz, 1H), 7.04 (d, J ) 4.0
Hz, 1H), 6.38 (d, J ) 4.1 Hz, 1H), 4.89-4.84 (m, 1H), 3.75 (s,
3H), 1.88-1.82 (m, 6H), 1.67-1.62 (m, 2H), and 1.32 (s, 6H).

1872 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 6
Fensome et al.
or test compounds are added in the presence of 1 nM progesterone.
The cells are incubated at 37 °C in a 5% CO2 humidified atmosphere
for 24 h.
treatment, they were allowed to rest for at least two cycles before
reassignment for the next treatment, except the vehicle-treated
group in which animals were rested for one cycle. The monkeys
were treated for 30 days (day 2–31; the first day of menses was
designated as day 1 of menstrual cycle). The compounds (in
2% Tween 80/0.5% methyl cellulose, 2 mL/kg) were given orally
once daily, and blood samples were collected by venipuncture
before the first dose and every Monday, Wednesday, and Friday
until next menses. Serum samples were collected and stored at
–80 °C until analyzed for progesterone levels.
Alkaline Phosphatase Enzyme Assay. At the end of treatment,
the medium is removed from the plate. Fifty microliters of 0.1 M
Tris-HCl, pH 9.8, containing 0.2% Triton ꢀ-100 is added to each
well. The plates are shaken in a titer plate shaker for 15 min. Then
150 µL of p-nitrophenyl phosphate (4 mM in 0.1 M Tris-HCl, pH
9.8) is added to each well. Optical density measurements are taken
at 5 min intervals for 30 min at a test wavelength of 405 nM. Data
is analyzed using JMP software (SAS Institute, Inc.) for both one-
way analysis of variance and non-4 linear dose response analysis
in both single dose and dose response studies.
Serum progesterone levels were measured by radioimmunoas-
say.36 The sensitivity of this assay was 2.5 pg/tube and the intra-
and interassay percent coefficient of variation at approximately the
IC50 value were 6.6% and 12.3%, respectively.
Progesterone Receptor Whole Cell Competition Binding
Assay. T47D Cell Culture. T47D cells are maintained in 10% RC
media at 37 °C in a 5% CO2 humidified atmosphere and needed to
be split twice weekly for proper response. Cells are plated in 10%
RC the day before the binding assay at 50 000 cells per well in the
white, clear bottom plates.
Acknowledgment. The authors wish to thank the Depart-
ment of Analytical Chemistry, Wyeth, for analytical data. We
also wish to thank our management, Drs. M Abou-Gharbia, R.
Magolda, and L. Freedman, for their continued support of this
work.
Binding Assay. Cells plated the day prior to the assay in white
clear bottom plates are used. A master compound plate is set up
containing control and test compounds at 20× final desired
concentration for the competition binding. Control compounds are
typically run 10-fold lower than this and include a 0 or vehicle
(50% DMSO/50% ethanol (v/v)) control well. A stock of 60 nM
3H-progesterone (Perkin-Elmer Life Science, NET-381, typically
around 102 Ci/mmol) (20×) is also prepared at a volume needed
of 10 µL per well. Media on cells is replaced with 180 µL of 5%
Supporting Information Available: Analytical HPLC and low-
resolution mass spectrometry data for compounds 12-21, 25-26,
27, 28–30, and 37–48. This material is available free of charge via
the Internet at http://pubs.acs.org.
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