Estrogen receptor ligands. Part 14: Application of novel antagonist side chains to existing platforms

Article abstract of DOI:10.1016/j.bmcl.2005.08.084

Two novel side chains which had previously been found to enhance antagonist activity in the dihydrobenzoxathiin SERM series were applied to three existing platforms. The novel side chains did not improve the antagonist activity of the existing platforms.

Full text of DOI:10.1016/j.bmcl.2005.08.084

Bioorganic & Medicinal Chemistry Letters 15 (2005) 5124–5128
Estrogen receptor ligands. Part 14: Application of novel
antagonist side chains to existing platforms
Timothy A. Blizzard,* Jerry D. Morgan, II, Wanda Chan, Elizabeth T. Birzin, Lee-Yuh Pai,
Edward C. Hayes, Carolyn A. DaSilva, Ralph T. Mosley, Yi Tien Yang, Susan P. Rohrer,
Frank DiNinno and Milton L. Hammond
Merck Research Laboratories, RY800-B116, PO Box 2000, Rahway, NJ 07065, USA
Received 29 July 2005; revised 22 August 2005; accepted 23 August 2005
Available online 3 October 2005
Abstract—Two novel side chains which had previously been found to enhance antagonist activity in the dihydrobenzoxathiin SERM
series were applied to three existing platforms. The novel side chains did not improve the antagonist activity of the existing
platforms.
Ó 2005 Elsevier Ltd. All rights reserved.
The importance of the selective estrogen receptor modu-
lators (SERMs) is well documented.1 The discovery of a
second ER subtype2 generated interest in the develop-
ment of subtype-selective SERMS.3 Previous publica-
tions from this laboratory have reported the discovery
of dihydrobenzoxathiins (e.g., 1) as a novel class of selec-
tive estrogen receptor alpha modulators (SERAMs).4a–e
OH
Since side chains A–C conferred such excellent antago-
nist properties on the dihydrobenzoxathiin platform,
we wondered if a similar effect would be observed with
other platforms or if A–C were platform-specific. Molec-
ular modeling of 1 versus raloxifene (2), bazedoxifene
(3), and lasofoxifene (4) in the hER-a binding site
showed considerable overlap of the side chains, suggest-
ing that side chains A–C might be of interest in these
other platforms (Fig. 1).5 While there have been a few
studies on side chain SAR of other platforms,6 we are
unaware of any systematic studies on the portability of
side chain SAR between platforms.
HO
S
O
N
O
1
We have also reported extensive studies on the side
chain SAR of this class.4d–g These studies resulted in
the discovery of two significantly improved antagonist
side chains, A and B,4f which were combined to afford
an optimized antagonist side chain C.4g
We therefore targeted analogs 2a,b, 3a,b, and 4a,b for
synthesis (Schemes 1–3 and Table 1). We chose the par-
tially optimized side chains A and B for these studies,
rather than the fully optimized side chain C, to afford
maximum insight into the applicability of our SAR to
other platforms. If substantial improvement was seen
in any platform with side chains A and B, then we would
prepare the corresponding analog with side chain C.
N
A
N
The raloxifene analogs 2a (R = A) and 2b (R = B) were
easily synthesized from fluoride 57 and the appropriate
side-chain alcohol4e using a modification of the Lilly
groupÕs raloxifene analog synthesis (Scheme 1).7,8
N
C
N
B
The bazedoxifene analogs 3a (R = A) and 3b (R = B)
were prepared from intermediate 6 and the appropriate
side-chain alcohol 74e using a modification of the Wyeth
groupÕs bazedoxifene synthesis (Scheme 2).9
Keywords: SERMs; SERAMs; Estrogen.
*
Corresponding author. Tel.: +1 732 594 6212; fax: +1 732 594
9556; e-mail: tim_blizzard@merck.com
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.08.084

T. A. Blizzard et al. / Bioorg. Med. Chem. Lett. 15 (2005) 5124–5128
5125
A
B
C
Figure 1. Molecular modeling of compound 1 (green) against models of (A) crystallographically determined raloxifene 2 (white); (B) bazedoxifene 3
(purple); and (C) lasofoxifene 4 (cyan) in the context of hER-a (pdb entry 1ERR).
F
RO
OBn
OBn
OBn
i - iii
O
O
i-iii
iv
Br
OMe
OH
Br
S
S
MeO
HO
MeO
MeO
5
2a,b
10
11
12
Scheme 1. Reagents: (i) ROH, NaH, DMF; (ii) HCl, ether; (iii) Br3,
CH2Cl2.
OH
OR
v-vi
vii-viii
ROH
CHO
i-iii
iv-vi
Cl
R = A or B
RO
RO
7
8
9
MeO
MeO
13
4a,b
RO
Scheme 3. Reagents: (i) Mg, THF, 30%; (ii) tetralone; (iii) pyridinium
tribromide, 76–100%; (iv) phenylboronic acid, (Ph3P)4Pd, THF, 71%;
(v) H2, Pd(OH)2/C, EtOH; (vi) chiral HPLC on Chiralcel AD column
eluted with heptane–isopropanol, 4:1; (vii) RCl prepared in situ from
ROH and SOCl2, THF, K2CO3, DMF; (viii) 2 N HCl, BBr3, Et2O,
CH2Cl2.
H
N
N
vii-viii
OBn
OH
BnO
HO
6
3a,b
Scheme 2. Reagents: (i) 2 N HCl, Et2O; (ii) SOCl2, THF; (iii) p-OH-
benzaldehyde, K2CO3, DMF (38% from 7); (iv) LiAlH4, Et2O, 91%;
(v) 2 N HCl, Et2O; (vi) SOCl2, THF; (vii) 9, NaH, DMF, 67%; (viii) Pd
(black) HCO2NH4; EtOH, EtOAc, H2O, 70%.
bazedoxifene, and lasofoxifene analogs all retained
potency comparable to or, at most, slightly better than
that of the corresponding parent.
Finally, the novel compounds were evaluated in an
immature rat uterine weight assay.13 This was the assay
where the largest impact was observed in the dihydro-
benzoxathiin series (compare 1a–c with 1). In the
raloxifene series, substitution of side chains A and B
for piperidine had no effect on the uterine activity of
2. In the bazedoxifene series, substitution of side chain
A for homopiperidine of 3 resulted in a slight increase
in uterine antagonism but substitution of side chain B
resulted in a decrease in antagonist activity. In the laso-
foxifene series, there was a slight increase in uterine
antagonism with a corresponding decrease in agonism
upon substitution of side chains A and B for pyrrolidine
in 4 but the effect is clearly not as dramatic as in the di-
hydrobenzoxathiin series.
Lasofoxifene analogs 4a (R = A) and 4b (R = B) were
prepared from commercial 10 and the appropriate
side-chain alcohol 74e using a modification of the Pfizer
groupÕs lasofoxifene synthesis (Scheme 3).10
Biodata for the novel analogs and SERAMs 1–1c are
summarized in Table 1. All compounds were evaluated
in an in vitro estrogen receptor (ER) binding assay.11
As we had observed in our previous studies with 1–1c,
substitution of the novel side chains A and B for the
side-chain of the parent compound did not have a signif-
icant effect on ER binding with any platform.
The compounds were also tested in an MCF-7 assay.12
In the dihydrobenzoxathiin series, the modified side
chains resulted in a modest potency enhancement in this
assay. However, a similar enhancement was not ob-
served with any of the other platforms. The raloxifene,
The lack of a dramatic effect on the uterine profile upon
incorporation of side chains A and B clearly indicates
that the side chain SAR of the dihydrobenzoxathiin

5126
T. A. Blizzard et al. / Bioorg. Med. Chem. Lett. 15 (2005) 5124–5128
Table 1. ER ligand binding data, rat uterine growth data, and MCF-7 data
OR
RO
RO
OH
HO
S
O
N
O
OH
OH
S
HO
HO
HO
OR
2
3
4
1
Compound
R
Human ER binding (IC50, nM)11
MCF-712 (IC50, nM)
Rat uterine activity13
hER-a
hER-b
b/a
% Ant.
72
% Ag.
34
1
2.6
3.0
1.3
1.3
1.8
0.6
0.211b
0.6
0.5
0.4
1.4
0.5
0.6
64
25
54
35
40
7
3.3
N
N
1a
1b
1c
2
161
45
0.9
0.3
0.2
0.8
1.0
0.2
0.4
0.3
0.1
0.1
0.08
0.07
106
101
123
81
À19
0
N
N
52
À29
24
12
N
2a
2b
3
3.1
2.611b
3.8
2.4
2.3
1.8
0.6
0.9
5
79
24
N
N
13
6
82
31
67
15
N
3a
3b
4
5
78
À2
20
N
N
6
61
1
74
26
N
4a
4b
1
89
10
N
N
1
81
18
SERAMs is not transferable to other platforms. Since
substantial improvement in the uterine profile was not
seen upon introduction of the improved antagonist side
chains A and B in the established platforms, we elected
not to prepare the corresponding analogs with the fully
optimized dihydrobenzoxathiin antagonist side chain C.

T. A. Blizzard et al. / Bioorg. Med. Chem. Lett. 15 (2005) 5124–5128
5127
S.; Schaeffer, J. M.; Hammond, M. L. Bioorg. Med. Chem.
Lett. 2004, 14, 2741; (d) Blizzard, T. A.; DiNinno, F.;
Morgan, J. D.; Chen, H. Y.; Wu, J. Y.; Gude, C.; Kim, S.;
Chan, W.; Birzin, E. T.; Yang, Y.; Pai, L.; Zhang, Z.;
Hayes, E. C.; DaSilva, C. A.; Tang, W.; Rohrer, S. P.;
Schaeffer, J. M.; Hammond, M. L. Bioorg. Med. Chem.
Lett. 2004, 14, 3861; (e) Blizzard, T. A.; DiNinno, F.;
Morgan, J. D., II; Wu, J. Y.; Chen, H. Y.; Kim, S.; Chan,
W.; Birzin, E. T.; Yang, Y.; Pai, L.; Zhang, Z.; Hayes, E.
C.; DaSilva, C. A.; Tang, W.; Rohrer, S. P.; Schaeffer, J.
M.; Hammond, M. L. Bioorg. Med. Chem. Lett. 2004, 14,
3865; (f) Blizzard, T. A.; DiNinno, F.; Morgan, J. D., II;
Chen, H. Y.; Wu, J. Y.; Kim, S.; Chan, W.; Birzin, E. T.;
Yang, Y.; Pai, L.; Fitzgerald, P. M. D.; Sharma, N.; Li,
Y.; Zhang, Z.; Hayes, E. C.; DaSilva, C. A.; Tang, W.;
Rohrer, S. P.; Schaeffer, J. M.; Hammond, M. L. Bioorg.
Med. Chem. Lett. 2005, 15, 107; (g) Blizzard, T. A.;
DiNinno, F.; Chen, H. Y.; Kim, S.; Wu, J. Y.; Chan, W.;
Birzin, E. T.; Yang, Y.; Pai, L.; Hayes, E. C.; DaSilva, C.
A.; Rohrer, S. P.; Schaeffer, J. M.; Hammond, M. L.
Bioorg. Med. Chem. Lett. 2005, 15, 3912.
In conclusion, we have demonstrated that the side chain
SAR of the dihydrobenzoxathiin SERAMs is not appli-
cable to existing platforms. Clearly, it will be necessary
to conduct SAR studies for each platform to determine
its optimum side chain. This concludes our study of the
dihydrobenzoxathiin side chain SAR.
Acknowledgments
The authors thank Dr. Derek Von Langen and Mr. Ste-
ven Fabian for the chiral prep HPLC purification of a
lasofoxifene intermediate. The authors also thank Ms.
Judith Pisano for helpful discussions.
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1
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11. (a) The IC50 values were generated in an estrogen receptor
ligand binding assay. This scintillation proximity assay
was conducted in NEN Basic Flashplates using tritiated
estradiol and full length recombinant human ERa and
ERb proteins. Compounds were evaluated in duplicate in
a single assay. In our experience, this assay provided IC50
values that are reproducible to within a factor of 2–3.
Dihydrobenzoxathiin 1 (n = l36) and estradiol (n > 100)
were tested in multiple assays; data reported in Table 1 are
an average of all determinations; (b) Data for 2b reflect a
20 h incubation prior to radioactive quantification; all
other data obtained with 3 h incubation.
4. (a) Kim, S.; Wu, J. Y.; Birzin, E. T.; Frisch, K.; Chan, W.;
Pai, L.; Yang, Y. T.; Mosley, R. T.; Fitzgerald, P. M. D.;
Sharma, N.; DiNinno, F.; Rohrer, S.; Schaeffer, J. M.;
Hammond, M. L. J. Med. Chem. 2004, 47, 2171; (b) Chen,
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W.; Yang, Y. T.; Colwell, L.; Li, S.; DiNinno, F.; Rohrer,
12. This is an in vitro MCF-7 breast cancer cell proliferation
assay adapted to a 96-well format. Cells are grown in
estrogen-depleted media for six days and then treated with
the test compound for seven days. To evaluate the

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T. A. Blizzard et al. / Bioorg. Med. Chem. Lett. 15 (2005) 5124–5128
antagonist activity of a test compound, this treatment
and antagonism in rat uterine tissue. Compounds are
dosed orally at 1 mpk. Agonism is reported as percent-
age of estradiol control; antagonism reported as per-
centage antagonism of estradiol; (b) Estradiol exhibited
100% agonism at 4 lg/kg; (c) Wakeling, A. E.; Valcaccia,
B.; Newboult, E.; Green, L. R. J. Steroid Biochem. 1984,
20, 111.
occurs in the presence of 0.003 nM (the EC70) estradiol.
The amount of cell growth is determined by measuring the
protein content of living cells and an IC50 for the test
compound is determined.
13. (a) The uterine weight assay is an in vivo assay based on
a published procedure13c that measures estrogen agonism