SAR studies of 6-(arylamino)-4,4-disubstituted-1-methyl-1,4-dihydro-benzo[d][1,3]oxazin-2-ones as progesterone receptor antagonists

Article abstract of DOI:10.1016/j.bmcl.2006.09.060

We previously disclosed that 6-aryl benzoxazin-2-ones were PR modulators. In a continuation of this work we examined the SAR of new 6-arylamino benzoxazinones and found the targets 1-25, with an extra amino linker between the pendent 6-aryl groups and ben

Full text of DOI:10.1016/j.bmcl.2006.09.060

Bioorganic & Medicinal Chemistry Letters 17 (2007) 189–192
SAR studies of 6-(arylamino)-4,4-disubstituted-1-methyl-1,4-
dihydro-benzo[d][1,3]oxazin-2-ones as progesterone
receptor antagonistsq
Jeffrey C. Kern,a Eugene A. Terefenko,a Andrew Fensome,a Ray Unwallla,a Jay Wrobel,a
Yuan Zhu,b Jeffrey Cohen,b Richard Winneker,b Zhiming Zhangb and Puwen Zhanga,*
aChemical and Screening Sciences, Wyeth Research, 500 Arcola Road, Collegeville, PA 19426, USA
bWomen’s Health and Musculoskeletal Biology, Wyeth Research, 500 Arcola Road, Collegeville, PA 19426, USA
Received 18 August 2006; revised 20 September 2006; accepted 21 September 2006
Available online 5 October 2006
Abstract—We previously disclosed that 6-aryl benzoxazin-2-ones were PR modulators. In a continuation of this work we examined
the SAR of new 6-arylamino benzoxazinones and found the targets 1–25, with an extra amino linker between the pendent 6-aryl
groups and benzoxazinone or benzoxazine-2-thione core, were PR antagonists. A series of compounds with substituents at the
1- and 4-positions as well as different 6-aryl groups were prepared and tested in the T47D cell alkaline phosphatase assay. Interest-
ingly, the SAR unveiled from the 6-arylamino benzoxazinones was quite different from those of their parent compounds. For
example, in contrast to the 6-aryl benzoxazinones, methyl substitution at the 1-position significantly increased the potency of
6-arylamino benzoxazinones. Several 6-arylamino benzoxazinones (e.g., 12, IC50 = 5.0 nM) had low nanomolar in vitro potency
as PR antagonists in the T47D cell alkaline phosphatase assay.
Ó 2006 Elsevier Ltd. All rights reserved.
Progesterone receptor (PR) is a member of the super-
family of ligand-dependent transcription factors.1 Pro-
gesterone is an endogenous hormone which plays an
important role in female reproduction. PR agonists have
been used widely in contraceptives and hormone thera-
py, often in combination with an estrogen. A selective
PR antagonist may be potentially used in female contra-
ception,2 and for the treatment of various gynecological
and obstetric diseases including hormone-dependent
cancers and non-malignant chronic conditions such as
fibroids and endometriosis.3,4 Clinically successful PR
antagonists, however, are limited and their therapeutic
potential has not yet been fully realized. Mifepristone,
the only marketed PR antagonist, demonstrated activity
at other steroidal receptors such as glucocorticoid (GR)
and androgen receptors and was nearly equipotent as an
antagonist for both PR and GR. This potentially limits
its chronic use.5 To search for more selective PR antag-
onists, a number of non-steroidal PR antagonists have
been investigated.6–9 Recently, we have disclosed three
series of novel PR antagonists including 6-aryl ben-
zimidazolones (e.g., 31), 5-aryl oxindoles (e.g., 32), and
6-aryl benzoxazinones (e.g., 33).10–12 Similar SAR
trends were observed for the benzimidazolone and oxin-
dole scaffolds. However, the PR functional activities of
the 6-aryl benzoxazinones were dependent on the nature
of the 6-aryl group and a few analogs showed modest
PR agonist activity in the T47D cell alkaline phospha-
tase assay.11 To further explore the SAR of 6-substitu-
tion from the benzoxazinone template, our strategy
was to place a linker between the 6-aryl groups and ben-
zoxazinone core. A number of oxygen, nitrogen, carbon,
and sulfur based linkers were evaluated. 6-Arylamino
benzoxazinones (1–25) were among the most potent
PR antagonists. The synthesis and in vitro SAR of a ser-
ies of novel 6-arylamino benzoxazinones as PR antago-
nists are the subject of this report.
Keywords: Progesterone receptor (PR); Non-steroidal PR modulator;
PR antagonists.
q
Presented in preliminary form at 230th ACS National Meeting,
Washington, DC, USA, August 28–September 1, 2005.
The preparation of target compounds 1–25 is shown in
Scheme 1. Addition of the appropriate Grignard reagent
to ketones 26 afforded amino carbinols 27. Ring closure
*
Corresponding author. Tel.: +1 484 865 3856; fax: +1 484 865
9398; e-mail: ZhangP@wyeth.com
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.09.060

190
J. C. Kern et al. / Bioorg. Med. Chem. Lett. 17 (2007) 189–192
R3 R2
O
alkaline phosphatase assay, the results are detailed in
Tables 1–4.
H
N
Ar
N
Cl
O
N
R1
O, S
As shown in Table 1, 6-arylamino benzoxazinones
methylated at the 1-position (2) resulted in about 4-fold
improvement in PR antagonist potency over their des-
methyl analogs 1. This is interesting since the NH moi-
ety at the 1-position has been demonstrated to play a
critical role in the PR potency for the previously report-
ed benzoxazinone scaffold (e.g., 6-(3-bromophenyl)-
1,4,4-trimethyl-1H-benzo[d][1,3]oxazin-2(4H)-one 33b
is more than 10-fold less potent than its des-methyl ana-
log 33a). This finding suggested that 6-arylamino ben-
zoxazinones and their parent compounds (e.g., 33)
may interact differently at the PR binding pocket.
N
H
1-25
31, IC50 = 73.1 nM
Cl
Br
O
O
N
N
O
H
R
33a, R = H, IC50 = 3.3 nM
33b, R = CH3, IC50 = 57.9 nM
32, IC50 = 56.0 nM
Interestingly, alkylation of the 1-position with a group
larger than the methyl group resulted in derivatives with
reduced PR antagonist potency (Table 1). The 1-methyl
benzoxazinone 3 exhibited moderate PR antagonist
potency with an IC50 = 54.4 nM. Increasing the size of
the 1-position substituent in compounds 4–7 caused a
17-fold or greater reduction in PR potency suggesting
a size limitation at this position.
of alcohols 27 with carbonyldiimidazole provided the
benzoxazinones 28. Treatment of the cyclic carbamates
28 with 70% nitric acid yielded exclusive nitration at
the 6-position resulting in 29. Alkylation of 29 with var-
ious alkyl iodides followed by reduction of the nitro
group afforded 1-alkylated intermediates 30 or direct
reduction of 29 gave the 1-unsubstituted 30. The ben-
zoxazinone target compounds 1–8 and 11–25 were ob-
tained by either coupling 30 to an appropriate boronic
acid via a copper (II) acetate mediated protocol or by
a palladium catalyzed reaction with an appropriate aryl
iodide. The benzoxazine-2-thiones 9 and 10 were readily
prepared by refluxing compounds 3 and 8 with Lawes-
son’s reagent in toluene.
Table 1. Inhibition of progesterone induced alkaline phosphatase
activity in T47D cells. SAR at the 1-position
F
H
N
X
O
O
R4
N
O
Among three series of PR antagonists we reported
recently,10–12 the 6-aryl benzoxazinone template (e.g.,
33) provided the most potent and selective PR antago-
nists as demonstrated in a number of in vitro and in vivo
models. In addition, the PR functional activities of the
6-aryl benzoxazinones were dependent on the nature
of the 6-aryl group.11 Thus, the benzoxazinone template
can provide, with a prudent choice of 6-aryl group, both
PR agonists and antagonists. We decided to place a link-
er between the 6-aryl moiety and benzoxazinone core
and examine the SAR of various linkers on the PR
potency and functional activity of target compounds.
From a handful of linkers that were screened, com-
pounds with an NH linker were among the most potent
PR antagonists. We therefore focused our effort on the
6-arylamino benzoxazinone template and prepared a
number of compounds with various substitution pat-
terns. These derivatives were evaluated in the T47D cell
N
O
H
R1
33c, R = Cl
33d, R = CN
Compound R1
R4
a
PR Alk. Phos. IC50 (nM)
1
2
H
20-Cl, 30-Cl
20-Cl, 30-Cl
40-Br
205.8
56.0
54.4
CH3
CH3
C2H5
3
4
40-Br
1000
1000
3000
3000
23.0
5
i-C3H7 40-Br
Phenyl 40-Br
Benzyl 40-Br
6
7
33c
33d
15.1
a 50% inhibitory concentration of tested compounds on 1 nM proges-
terone induced alkaline phosphatase activity in the human T47D
breast carcinoma cell line. Values represent the average of at least
duplicate determinations. The standard deviation for the assay was
20% of the mean or less.
6-Arylamino benzoxazin-2-ones and benzoxazine-2-thiones
R3 R2
O
R3 R2
O
R3 R2
O
H
O
R3 R2
OH
R
N
Ar
R2
f or g
h
b
c-e
a
N
R1
O
N
R1
X
N
O
NH2
26
NH2
27
H
28
29, R = NO2
30, R = NH2
1-8, 11-25: X = O
9, 10: X = S
Scheme 1. Reagents and conditions: (a) R3MGBr, THF, 0 °C, N2; (b) CDI, THF, rt, (c) HNO3, H2SO4, AcOH; (d) NaH, THF, 0 °C, N2, then R1X;
(e) NaBH4, MeOH, Pd/C, 0 °C, N2; (f) Cu(OAC)2, Et3N, aryl boronic acid, CH2Cl2, rt; (g) Pd2(dba)3, Cs2CO3, BINAP, 18-C-6, aryl iodide, THF,
40 °C; (h) Lawesson’s reagent, toluene, reflux, N2.

J. C. Kern et al. / Bioorg. Med. Chem. Lett. 17 (2007) 189–192
191
Table 2. Inhibition of progesterone induced alkaline phosphatase
activity in T47D cells. SAR at the 2-position
Table 4. Inhibition of progesterone induced alkaline phosphatase
activity in T47D cells. SAR of the 6-aryl group
R3 R2
H
N
H
O
N
R4
O
R4
N
X
N
O
CH3
CH3
Compound R2/R3 R4
X
a
PR Alk. Phos. IC50 (nM)
a
PR Alk. Phos. IC50 (nM)
Compound
R4
3
8
CH3
C2H5 30-F, 40-F
CH3
40-Br
C2H5 30-F, 40-F
40-Br
O
O
S
54.4
27.4
30.0
42.2
23.0
15.1
12
13
14
19
20
21
22
23
24
25
33c
33d
20-Cl, 30-Cl
30-Cl, 40-Cl
30-Cl
30-F, 40-F
30-Cl, 40-F
30-F, 40-CN
30-CN, 40-F
30-NO2, 50-F
30-Cl, 50-Cl
30-CN, 50-F
5.0
29.8
13.1
27.4
9.0
9
10
33c
33d
S
37.1
9.2
a 50% inhibitory concentration of tested compounds on 1 nM proges-
terone induced alkaline phosphatase activity in the human T47D
breast carcinoma cell line. Values represent the average of at least
duplicate determinations. The standard deviation for the assay was
20% of the mean or less.
8.4
8.1
14.5
23.0
15.1
a 50% inhibitory concentration of tested compounds on 1 nM proges-
terone induced alkaline phosphatase activity in the human T47D
breast carcinoma cell line. Values represent the average of at least
duplicate determinations. The standard deviation for the assay was
20% of the mean or less.
Table 3. Inhibition of progesterone induced alkaline phosphatase
activity in T47D cells. SAR at the 4-position
R3 R2
H
N
O
R4
N
O
As shown in Table 3, 6-arylamino benzoxazinones
substituted at the 4-position by diethyl (12 and 13)
showed modest improvement (3- to 10-fold) in PR
antagonist potency over their dimethyl substituted coun-
terparts (2 and 11). The 4-position was further explored
by replacing one of the ethyl groups with various aryl
moieties (15–17) shown in Table 3. In these examples
no further improvement was observed. Instead, racemic
compounds 15–17 were at least 2-fold less potent than
their parent compound 14. Replacement of both ethyl
groups of compound 14 with the 40-chlorophenyl group
(18) resulted in a dramatic loss in antagonist potency.
CH3
Compound R2
R3
R4
PR Alk.
Phos. IC50
(nM)
a
2
11
12
13
14
15
16
17
18
33c
33d
CH3
CH3
CH3
CH3
20-Cl, 30-Cl
30-Cl, 40-Cl
20-Cl, 30-Cl
30-Cl, 40-Cl
30-Cl
30-Cl
30-Cl
56.0
100.0
5.0
C2H5
C2H5
C2H5
C2H5
C2H5
C2H5
C2H5
C2H5
29.8
13.1
32.2
66.1
71.8
1000
23.0
15.1
C2H5
Thien-2-yl
Phenyl
4-Cl-phenyl 30-Cl
4-Cl-phenyl 4-Cl-phenyl 30-Cl
Using the optimized 4,4-diethyl-1-methylbenzoxazinone
core, a number of 6-aryl groups containing various elec-
tron-withdrawing substituents were examined as depict-
ed in Table 4. Overall, these analogs had good to
excellent PR antagonist potency (IC50 = 5.0–37.4 nM).
Compounds 20 and 22 had IC50 of 9.0 and 9.2 nM
and were more potent than their corresponding 6-aryl
benzoxazinone analogs11 33c and 33d (IC50 = 23.0 and
15.1 nM, respectively).
a 50% inhibitory concentration of tested compounds on 1 nM proges-
terone induced alkaline phosphatase activity in the human T47D
breast carcinoma cell line. Values represent the average of at least
duplicate determinations. The standard deviation for the assay was
20% of the mean or less.
In summary, we have demonstrated that 6-arylamino
benzoxazinones are PR antagonists, which have a dif-
ferent SAR when compared to that of parent 6-aryl
benzoxazinones. In contrast to their parent com-
pounds, a methyl substitution at the 1-position signif-
icantly increased the potency. Diethyl substitution at
the 4-position was found to be optimal. Several 6-
arylamino benzoxazinones (e.g., 12, IC50 = 5.0 nM)
had low nanomolar in vitro potency and were potent
PR antagonists in the T47D cell alkaline phosphatase
assay.
Substitution of a thio-carbonyl group at the 2-position for
the carbonyl moiety, in general, switched PR antagonists
to agonists for the oxindole (e.g., 32) and benzoxazinone
(e.g., 33) templates.13,14 However, as illustrated in Table
2, this functional activity switch did not take place in
the 6-arylamino benzoxazinone scaffold. The benzox-
azine-2-thiones 9 and 10 remained as PR antagonists
(IC50 = 30.0, 42.2 nM) and had similar potency as their
parent carbonyl compounds 3 and 8 (IC50 = 54.4,
27.4 nM).

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J. C. Kern et al. / Bioorg. Med. Chem. Lett. 17 (2007) 189–192
7. Hamann, L. G.; Winn, D. T.; Pooley, C. L. F.; Tegley, C.
Acknowledgments
M.; West, S. J.; Farmer, L. J.; Zhi, L.; Edwards, J. P.;
Marschke, K. B.; Mais, D. E.; Goldman, M. E.; Jones, T.
K. Bioorg. Med. Chem. Lett. 1998, 8, 2731.
We thank Drs. Ron Magolda and Magid Abou-Ghar-
bia for support and encouragement, Dr. Eugene
Trybulski for scientific suggestions, and the assistance
of Department of Analytical Chemistry for analytical
data.
8. Kurihara, K.; Tanabe, K.; Yamamoto, Y.; Shinei, R.; Ajito,
K.; Okonogi, T. Bioorg. Med. Chem. Lett. 1999, 9, 1837.
9. Zhi, L.; Tegley, C. M.; Pio, B.; West, S. J.; Marschke, K.
B.; Mais, D. E.; Jones, T. K. Bioorg. Med. Chem. Lett.
2000, 10, 415.
10. Zhang, P.; Terefenko, E. A.; Wrobel, J.; Zhang, Z.; Zhu,
Y.; Cohen, J.; Marschke, K. B.; Mais, D. Bioorg. Med.
Chem. Lett. 2001, 11, 2747.
References and notes
11. Zhang, P.; Terefenko, E. A.; Fensome, A.; Wrobel, J.;
Winneker, R.; Lundeen, S.; Marschke, K. B.; Zhang, Z.
J. Med. Chem. 2002, 45, 4379.
12. Fensome, A.; Bender, R.; Cohen, J.; Collins, M. A.;
Mackner, V. A.; Miller, L. L.; Ullrich, J. W.; Winneker,
R.; Wrobel, J.; Zhang, P.; Zhang, Z.; Zhu, Y. Bioorg.
Med. Chem. Lett. 2002, 12, 3487.
13. Zhang, P.; Terefenko, E. A.; Fensome, A.; Wrobel, J.;
Winneker, R.; Zhang, Z. Bioorg. Med. Chem. Lett. 2003,
13, 1313.
14. Fensome, A.; Koko, M.; Wrobel, J.; Zhang, P.; Zhang, Z.;
Cohen, J.; Lundeen, S.; Rudnick, R.; Zhu, Y.; Winneker,
R. Bioorg. Med. Chem. Lett. 2003, 13, 1316.
1. Mangelsdorf, D. J.; Thummel, C.; Beato, M.; Herrlich, P.;
Schuetz, G.; Umesono, K.; Blumberg, B.; Kastner, P.;
Mark, M. Cell (Cambridge, MA) 1995, 83, 835.
2. Brown, A.; Cheng, L.; Lin, S.; Baird, D. T. J. Clin.
Endocrinol. Metabol. 2002, 87, 63.
3. Murphy, A. A.; Kettel, L. M.; Morales, A. J.; Roberts, V.
J.; Yen, S. S. J. Clin. Endocrinol. Metabol. 1993, 76, 513.
4. Kettel, L. M.; Murphy, A. A.; Morales, A. J.; Ulmann, A.;
Baulieu, E. E.; Yen, S. S. Fertility Sterility 1996, 65, 23.
5. Brogden, R. N.; Goa, K. L.; Faulds, D. Drugs 1993, 45,
384.
6. Combs, D. W.; Reese, K.; Phillips, A. J. Med. Chem. 1995,
38, 4878.