Synthesis and evaluation of hexahydrochrysene and tetrahydrobenzofluorene ligands for the estrogen receptor

Article abstract of DOI:10.1016/S0960-894X(01)00189-5

To prepare novel estrogen receptor (ER) ligands, we have developed a facile approach to substituted hexahydrochrysene and tetrahydrobenzo[a]fluorene systems. Substituents, including basic side chains, were added to these systems, and their binding affinity to ERα and ERβ, and in some cases their transcriptional activity were evaluated.

Full text of DOI:10.1016/S0960-894X(01)00189-5

Bioorganic & Medicinal Chemistry Letters 11 (2001) 1281–1284
Synthesis and Evaluation of Hexahydrochrysene and
Tetrahydrobenzofluorene Ligands for the Estrogen Receptor
Rosanna Tedesco, Michael K. Youngman, Scott R. Wilson and
John A. Katzenellenbogen*
Department of Chemistry, University of Illinois, 600 S. Mathews Ave., Urbana, IL 61801, USA
Received 23 January 2001; accepted 12 March 2001
Abstract—To prepare novel estrogen receptor (ER) ligands, we have developed a facile approach to substituted hexahydrochrysene
and tetrahydrobenzo[a]fluorene systems. Substituents, including basic side chains, were added to these systems, and their binding
affinity to ERa and ERb, and in some cases their transcriptional activity were evaluated. # 2001 ElsevierScience Ltd. All rights
reserved.
Selective estrogen receptor modulators (SERMs)1 are
estrogens that show a distinct pattern of tissue selective
action, possibly based on theirselective binding to the
two ER subtypes, ERa and ERb,2 ortheirability to
induce distinct receptor conformations.3 We have stud-
ied certain chrysene derivatives as part of an effort to
understand how ligand structural features determine ER
subtype selectivity and impart SERM-like profiles. The
5,6,11,12-tetrahydrochrysenes (THCs), first developed
as fluorescent ER ligands (1),4 proved to be particularly
interesting, because the cis-isomers, such as 2, display
remarkable ER subtype efficacy selectivity, being ago-
nists on ERa and antagonists on ERb.5
unstable, and were not studied.) We also introduced a
piperidinyl-ethoxyphenyl substituent, referred as a basic
side chain (BSC), into both the HHCs and TBHF.
Despite their overall similarity, these two ligand series
have considerably different shapes, resulting in a differ-
ent topological display of substituents around the rigid
ligand core. This has a significant effect on the ER
binding of the HHCs and THBFs.
Synthesis of Ketones 8ab, Key Intermediates for the
HHC and THBF Series
The tetracyclic ketones 8a and 8b proved to be versatile
intermediates for the synthesis of substituted HHCs and
THBFs, respectively. They were readily prepared
through their acyclic ketone precursors, 7a and 7b
(Scheme 1). A Shapiro reaction on tosylhydrazone 6
generated a vinyllithium intermediate that was trapped
with tributyltin chloride.6 Stille coupling7 of the result-
ing vinyl stannane with m-methoxyphenylacetyl chlo-
ride or m-anisoyl chloride gave the acyclic ketones 7a
and 7b, respectively, in good yield. These were cyclized
with methanesulfonic acid (MsOH) to give the desired
cyclic ketones 8a and 8b, as well as minoramounts of
To examine these systems further, we now describe the
preparation of saturated analogues of the THCs, the
hexahydrochrysenes (HHC, 3), as well as saturated
analogues of the related benzo[a]fluorene core, namely
tetrahydrobenzo[a]fluorenes (THBF, 5). (The dihy-
drobenzo[a]fluorenes (DHBF, 4) proved to be extremely
*Corresponding author. Tel.:+1-217-333-6310; fax; +1-217-333-
7325; e-mail: jkatzene@uiuc.edu
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00189-5

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R. Tedesco et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1281–1284
the isomeric products derived from attack ortho to the
methoxy group (9). In the HHC series, ketone 8b also
provided the spiro compound 10.
in the plane of the benzocyclopentane bicyclic portion,
whereas the cis-fused benzocyclohexane ring portion is
disposed in nearly a perpendicular arrangement with
respect to the benzocyclopentane. The basic side-chain
substituted THBF 21 was synthesized by coupling of 20
with piperidine ethanol and removal of the methoxy-
protecting groups.10
Underall of the cyclization conditions we examined, the
majorketone products 8ab in both the HHC and THBF
series had a cis-ring junction. This was confirmed by X-
ray crystallography for the THBF derivative, and was
ascertained by 1H NMR in the HHC case. Furthermore,
in the lattercase, erduction, deoxygenation and deme-
thylation8 of ketone 8b (Scheme 2) yielded material that
was identical to the known cis-hexahydrochrysene diol
12.9
Synthesis of Substituted Hexahydrochrysene Systems
Attempts to add Grignard reagent 18 orits ceirum
analogue to ketone 8b gave only fully aromatized pro-
ducts. However, Suzuki coupling11 of boronic acid 23
with the mixture of enol triflates 22, derived from
ketone 8b, gave regioisomers 24 (69%) and 25 (18%)
(Scheme 5). While reduction of 25 gave a mixture of
isomeric products, reduction of 24 with Et3SiH/TFA
Synthesis of Substituted Tetrahydrobenzo[a]fluorene
Systems
.
Ketone 8a was deprotected to the bisphenol 13 and also
converted to the THBF-2,8-diol 14 by deoxygenation
and deprotection. Addition of EtMgBr to ketone 8a
gave 15 as a single epimer of undetermined configura-
tion. After deoxygenation, a 2:1 mixture of diastereo-
isomers (16ab) was obtained, which could be separated
afterdeprotection ( 17ab) (Scheme 3). We were unable to
and BF3 Et2O gave primarily the saturated system 26.
The TBS group, partially removed during this reduc-
tion, was fully cleaved by treating the crude reaction
product with TBAF. The basic side chain was added to
1
assign the 17ab stereoisomers by H NMR analysis.
To add a basic side-chain substituent, ketone 8a was
treated with the Grignard reagent 18 to afford 19, which
was deoxygenated by treatment with Et3SiH and
.
BF3 Et2O followed by the addition of TFA. Monophenol
20 was obtained afterselective ermoval of the TBS
protecting group (Scheme 4).
We could not assign the relative configuration of 20 by
1H NMR; however, X-ray analysis established the cis–
cis configuration, in which all of the protons are on the
convex face of the molecule. The phenyl substituent in
this THBF system adopts an orientation that is almost
Scheme 3.
Scheme 1.
Scheme 2.
Scheme 4.

R. Tedesco et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1281–1284
1283
the free phenol 26 by standard methods,10 and final
deprotection was accomplished using AlBr3/EtSH to
afford 27.
the tetrahydrochrysene series,5 but very different from
the high ERa binding selectivities of a number of triaryl
pyrazoles we have investigated.15
The cis–cis configuration, initially assigned from 1H
NMR coupling constants, was confirmed by X-ray
crystallography. The conformation of this HHC system
is quite different from that of the THBF system 21
described above: the ring with the phenyl substituent
adopts a half chairconfomr ation, placing the phenyl
substituent in an axial-like position where it avoids
nonbonding interaction with the other aryl groups, and
the rings of the decalin system are more nearly coplanar
than they were in the THBF one.
The HHC and THBF analogues having the piperidinyl-
ethoxyphenyl basic side-chain substituent, 21 and 27,
present an interesting contrast. The THBF analogue 21
has much higheraffinity than the HHC analogue 27,
and both have a significant ERa affinity selectivity. It is
not easy to account forthese affinity and selectivity dif-
ferences, because they are probably related to differ-
ences in how the substituted and unsubstituted
compounds interact with various residues in the ligand-
binding pockets of ERa and ERb.
The relative binding affinity (RBA, E2=100%) of the
compounds prepared above was determined in a com-
petitive radiometric binding assay, using purified, full-
length human ERa and ERb (PanVera).12 The results of
these assays are summarized in Table 1, which also
includes RBA values on some related compounds whose
preparation, by other routes, is described elsewhere.13 A
few of the most interesting compounds were also tested
in cell-based reporter gene transcription assays.14
We assayed the transcriptional activity of three ligands
(lacking the BSC) that showed both good affinity and
had higherER b binding affinity selectivity, namely 12,
trans-HHC13 and the cis-THBF 14; none of them
showed a significant preferential potency for either ER
subtype (data not shown). The results of transcription
activation assays with the two BSC compounds are
shown in Figure 1. The HHC-derivative 27 (Fig. 1A)
acts as a mixed agonist–antagonist through ERa and as
an antagonist through ERb. By contrast, the THBF
derivative 21 behaves as an antagonist on both recep-
tors (Fig. 1B). The higher antagonistic character of the
THBF compound 21 could be related to the conforma-
tion of the BSC relative to the two ring systems. As one
might expect, changes in this orientation could preclude
orfavorintearctions between the basic side chain and
the key residues in the receptor (e.g., ER-Asp354).16,17
Apart from the compounds bearing a BSC, 21 and 27,
all of the derivatives have some selectivity for binding to
ERb. Both cis and trans unsubstituted HHCs have
higheraffinity than theirunsubstituted THC parent ( 1,
R=R0=H, X=OH), though theirER b affinity selec-
tivity is dramatically different. A diethyl HHC (3,
R=R0=Et), of which only the trans–cis–trans isomer
was available,13 binds much less well than its THC par-
ent (2), but has the highest ERb affinity selectivity of all
the compounds we have investigated here. In the THBF
series, the ketone 13 has very low affinity, but the
unsubstituted system 14 and especially the two ethyl
epimers 17ab (of unassigned configuration) are very
high affinity ligands, having ERb affinities substantially
greater than that of estradiol, with one epimer (17b)
having significant ERb affinity selectivity. The moderate
ERb binding selectivity of these compounds is reminis-
cent of a numberof compounds we have descirbed in
We have developed effective synthetic approaches to the
hexahydrochrysene (HHC) system, long known as
estrogen receptor ligands, and to the related members of
tetrahydrobenzo[a]fluorene (THBF) system. Our
approach enables the addition of an alkyl or phenyl
substituent (the latterof which was adorned with a basic
side chain), and it produced compounds having pre-
dominantly the cis configuration at the internal ring
fusion. Some of the unsubstituted orethyl-substituted
systems showed moderate ERb binding affinity selectiv-
ity, but had essentially no potency and/orefficacy selec-
tivity in cell-based assays. The HHC and THBF systems
substituted with a basic side chain were antagonists on
Table 1. ERa and ERb binding affinities and selectivities
Compound
Relative binding affinity (RBA %)a
ERa
ERb
b/a
1
2
12
R,R0=H; X=OH5
3.0
23
11
14
1.0
0.53
8.7
150
48
6.5
144
71
130
16
2.0
63
300
280
10
2
6
6.7
8.9
16
3.8
7.2
2.0
5.9
0.26
0.06
di-Et-THC5
cis-HHC
trans-HHC13
R,R0=Et (t,c,t)13
THBF ketone
THBF
Et-THBF
Et-THBF
BSCb-THBF
BSCb-HHC
3
13
14
17a
17b
21
27
38
1.1
0.06
aRelative binding affinity (RBA) values, where estradiol=100%.
bBSC=basic side chain.
Scheme 5.

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R. Tedesco et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1281–1284
Figure 1. Transcriptional activity of 27 (left) and 21 (right) through ERa and ERb in human endometrial carcinoma (HEC-1) cells. All values are
relative to the response to 1 nM estradiol (100%). For methods, see refs 5 and 14.
Hamilton, P. T.; McDonnell, D. P.; Fowlkes, D. M. Proc.
both ERa and ERb, with the THBF derivative display-
ing a more complete antagonism and higher potency
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than the HHC derivative.
A synthesis of the THBF derivative 21 has recently been
described in a patent, together with an indication of its
utility for the treatment of osteoporosis and hyperlipid-
emia.18 The THBF used in that report was a mixture of
isomers, and the relative configuration of the compo-
nents of the mixture was not specified. The interesting
biological profile of this THBF derivative, together with
the antagonist character that we have observed in our
THBF and HHC basic side-chain derivatives, suggests
that it would be promising to study further the indivi-
dual diastereomers of these systems, because the geo-
metric relationship between the BSC substituent and the
ligand core can be varied dramatically.
8. Desmaele, D.; Mekouar, K.; d’Angelo, J. J. Org. Chem.
1997, 62, 3890.
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D. L.; Rowley, E. R.; Shetler, P. K.; Short, L. L.; Venugopa-
lan, M.; Yang, N. N.; Sato, M.; Glasebrook, A. L.; Bryant,
H. U. J. Med. Chem. 1998, 41, 1272.
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Acknowledgements
14. Sun, J.; Meyers, M. J.; Fink, B. E.; Rajendran, R.; Kat-
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This work was supported through research grants from
the NIH (5R37 DK15556 and CA18119) and instru-
mentation grants from the NIH and NSF. We are
grateful to Kathryn Carlson, Jun Sun and Benita Kat-
zenellenbogen forbiological assays.
16. Brzozowski, A. M.; Pike, A. C.; Dauter, Z.; Hub-
¨
bard, R. E.; Bonn, T.; Engstrom, O.; Ohman, L.; Greene,
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