Phenylcyclohexene and phenylcyclohexadine substituted compounds having retinoid antagonist activity

Article abstract of DOI:10.1016/S0960-894X(01)00058-0

Retinoids are natural and synthetic analoques of the hormone retinoic acid. Systemic retinoid agonist therapy is usually associated with toxic side effects, such as mucocutaneos toxicity, which may be alleviated by the use of topical retinoid antagonists. We report the synthesis and biological activity of a new series of potent, RAR-specific antogonists substituted with phenylcyclohexene and phenylcyclohexadiene groups.

Full text of DOI:10.1016/S0960-894X(01)00058-0

Bioorganic & Medicinal Chemistry Letters 11 (2001) 765±768
Phenylcyclohexene and Phenylcyclohexadiene Substituted
Compounds Having Retinoid Antagonist Activity
Richard L. Beard,a,* Elliott S. Klein,b Andrew M. Standeven,b Maria Escobarb
and Roshantha A. S. Chandraratnaa,b,
*
aDepartment of Chemistry, Retinoid Research, Allergan Incorporated, Irvine, CA 92623-9534, USA
bDepartment of Biology, Retinoid Research, Allergan Incorporated, Irvine, CA 92623-9534, USA
Received 14 November 2000; accepted 15 January 2001
AbstractÐRetinoids are natural and synthetic analogues of the hormone retinoic acid. Systemic retinoid agonist therapy is usually
associated with toxic side e€ects, such as mucocutaneous toxicity, which may be alleviated by the use of topical retinoid antago-
nists. We report the synthesis and biological activity of a new series of potent, RAR-speci®c antagonists substituted with phenyl-
cyclohexene and phenylcyclohexadiene groups. # 2001 Elsevier Science Ltd. All rights reserved.
Retinoids are natural and synthetic analogues of the
hormone retinoic acid.1 Retinoids are currently being
investigated clinically as drugs in several areas, includ-
ing dermatology and oncology. The therapeutic use of
retinoid agonists as drugs is limited because they elicit a
number of toxic side e€ects, such as teratogenicity,
headache, and mucocutaneous toxicity.2 Retinoid
antagonists may be useful to counteract some of the
adverse e€ects associated with retinoid therapy. For
example, retinoid antagonists may be used as a topical
treatment for the mucocutaneous toxicity caused by
13-cis-retinoic acid (AccutaneTM), a systemic retinoid
drug used to treat nodulocystic acne.
heterodimers are e€ectively activated by RAR-speci®c
agonists, but not by RXR-speci®c agonists.5 Depending
on the nature of the bound ligand, the receptors
undergo distinct conformational changes that allow
them to bind to co-activator6 or co-repressor7 proteins
that either activate or repress gene transcription,
respectively.
Retinoids belong to the steroid-thyroid-retinoid super-
family of hormones that induce biological responses by
binding to and activating nuclear receptors. Retinoids
are natural and synthetic ligands that bind to the six
known retinoid receptors, the retinoic acid receptors-
RARa, b, and gÀand the retinoid X receptorsÐRXRa,
b, and g. The natural physiological ligand for the RARs
is all-trans-retinoic acid (ATRA), which binds with high
anity to each of the RAR subtypes but does not bind
to the RXRs.3 The putative hormone for the RXRs is 9-
cis-retinoic acid, which binds to both RXRs and RARs
with high anity.4 In biological systems, RARs form
functional heterodimers with RXRs. These RAR±RXR
Recently we reported on the identi®cation of AGN
193109, a highly potent and selective RAR antagonist
and inverse agonist.8 We have also reported on several
novel structural classes of RAR selective antagonists,
which are analogues of AGN 193109.9 These com-
pounds are similar to one another in that each of them is a
highly rigid structure substituted with an aryl group in the
1-position (ATRA numbering). We wished to examine if
*Corresponding author. Fax: +1-714-246-4209;
e-mail: chandraratna_rosh@allergan.com
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00058-0

766
R. L. Beard et al. / Bioorg. Med. Chem. Lett. 11 (2001) 765±768
1-phenyl substituents would impart antagonist and/or
inverse agonist activity on a series of more ¯exible reti-
noid analogues, such as AGN 190121, a potent RAR
agonist.10 Thus, we synthesized two series of new com-
pounds that have structural features common to both
AGN 193109 and AGN 190121. The ®rst series includes
compounds 1±3, which possess phenyl groups in place
of the 18-methyl group of AGN 190121. The second
series, which includes compounds 4 and 5, have a gem-
dimethyl group at the 4-position and a phenyl group at
the 1-position and lack the 18-methyl group. We will
show that these compounds act as potent antagonists of
the potent RAR agonist, (E)-4-[2-(5,6,7,8-tetrahydro-
5,5,8,8-tetramethyl-2-naphthalenyl)propen-1-yl]benzoic
acid (TTNPB)11 in receptor transactivation assays.
Moreover, in a hairless mouse model of retinoid
induced topical irritation,12 compound 1 was able to
reverse the topical irritation caused by administration of
TTNPB.
catalysis to produce the desired 2-phenyl-6,6-dimethyl-
1-cyclohexene compounds as benzoate esters, which
were hydrolyzed to give the carboxylic acids
(R=methyl), 2 (R=ethyl), and 3 (R=tert-butyl) in
1
70%, 76%, and 89% yield, respectively.
A synthesis of the cyclohexadiene analogues used in this
study is illustrated in Scheme 2. The starting material,
4,4-dimethyl-2-cyclohexen-1-one (10), was iodinated in
the 2-position by following the procedure of Sha and
Huang.14 This iodide and (E)-1-(tri-n-butylstannyl)-2-
(trimethylsilyl)ethylene were reacted with a palladium
catalyst and the resulting vinylsilane was iodinated to
produce vinyl iodide 11 in 24% overall yield from 10.
Compound 11 and ethyl 4-ethynylbenzoate underwent
a Heck reaction to produce benzoate 12 in 73%
yield. Treatment of cyclohexenone 12 with sodium bis-
(trimethylsilyl)amide and N-phenyltri¯uoromethane-
sulfonimide gave the cyclohexadienyl tri¯ate 13 in 65%
yield. The tri¯ate 13 was reacted separately with 4-
tolylzinc bromide and 4-(tert-butyl)phenylzinc bromide
in the presence of palladium(0) to give the expected C1-
phenyl substituted derivatives, which were subsequently
hydrolyzed to give the corresponding carboxylic acids, 4
(R=methyl) and 5 (R=tert-butyl), in 81% and 53%
yield, respectively.
The analogues used in this study were obtained as fol-
lows.13 The cyclohexene derivatives were prepared from
2-iodo-3-methyl-2-cyclohexen-1-one (6)14 as shown in
Scheme 1. Palladium catalyzed coupling reaction of
6 with (E)-1-(tri-n-butylstannyl)-2-(trimethylsilyl)ethyl-
ene15 and subsequent iododesilation reaction gave the
(E)-vinyl iodide 7 in 30% yield. Compound 7 underwent
a Heck reaction with ethyl 4-ethynylbenzoate15 followed
by Michael reaction of the resulting cyclohexenone with
methyl cuprate to produce the 3,3-dimethylcyclohex-
anone derivative, compound 8, in 42% yield. The
ketone was converted into an enol tri¯ate, compound 9,
by treatment with tri¯ic anhydride and a hindered base.
This reaction also produced the other enol tri¯ate
regioisomer, which was separated by ¯ash column
chromatography, thereby lowering the yield of 9 to
42%. The tri¯ate was reacted with three separate arylzinc
reagents, 4-tolylzinc bromide, 4-ethylphenylzinc bro-
mide, 4-tert-butylphenylzinc bromide, under palladium
These compounds were tested in RAR binding and
transactivation assays as previously described,16 and the
results are summarized in Table 1. None of these com-
pounds exhibited any binding or transactivation activity
at the RXRs (data not shown). Although these com-
pounds do not activate any of the RAR subtypes, they
bind e€ectively to all three. With the exception of com-
pound 4, these compounds bind with highest anity to
the RARb subtype, which is consistent with the binding
properties of the RAR agonist, AGN 190121. These
compounds also have a preference for binding to
the RARg subtype relative to RARa. The 4-tolyl and
4-ethylphenyl substituted cyclohexene derivatives, com-
pounds 1 and 2, respectively, have higher binding
Scheme 1. Synthesis of 2-phenyl-6,6-dimethyl-1-cyclohexene retinoid
antagonists. (a) Bu3SnCHCHSiMe3, Pd(PPh3)4; (b) I2, THF, 30%
(two steps); (c) 4-EtO2CPhCCH, PdCl2(PPh3)2, CuI, Et3N, 50 ꢀC; (d)
Me2CuLi, TMSCl, HMPA; H3O+, 42% (two steps); (e) Tf2O, 2,6-di-
tert-butyl-4-methylpyridine, 42%; (f) ArZnBr, Pd(PPh3)4; (g) NaOH,
EtOH; H+, 70±89% (two steps).
Scheme 2. Synthesis of 6-phenyl-3,3-dimethyl-1,5-cyclohexadiene reti-
noid antagonists. (a) I2, TMSN3, pyridine, 85%; (b) Bu3SnCHCH-
SiMe3, Pd(PPh3)4, 69%; (c) I2, THF, 31%; (d) 4-EtO2CPhCCH,
PdCl2(PPh3)2, CuI, Et3N, 40 ꢀC, 73%; (e) NaHMDS; Tf2NPh, 65%;
(f) ArZnBr, Pd(PPh3)4; (g) NaOH, EtOH; H+, 53±81% (two steps).

R. L. Beard et al. / Bioorg. Med. Chem. Lett. 11 (2001) 765±768
767
anity to each of the RAR subtypes than the corre-
sponding 4-tert-butylphenyl analogue, 3. Interestingly,
the 4-tolyl substituted cyclohexadiene 4 binds equally
well to RARb and RARg, but the 4-tert-butylphenyl
substituted cyclohexadiene 5 binds exclusively to RARb
at the doses tested. Thus, in the cyclohexadiene series,
structural changes in this position a€ect ligand binding
to RARa and RARg more than to RARb. This trend is
also observed with dihydronaphthtalene substituted
antagonists such as AGN 193109.17
be desirable to inhibit the mucocutaneous toxicity pro-
duced by a systemically applied retinoid agonist. This
toxicity is generally limited to small areas around the
nose and on the lips. Thus, an antagonist applied topi-
cally to this relatively small area would have only mini-
mal absorption and should not interfere with the overall
bene®cial e€ects of the retinoid agonist. It is known that
RARg is the most abundant receptor in the skin,18 and
convincing evidence exists that the skin irritation
induced by retinoids is RARg mediated.19 Thus, the
RARg selectivity displayed by these compounds in the
inhibition of TTNPB-induced transactivation suggests
that they may be excellent candidates for the topical
treatment of the mucocutaneous toxicity that is asso-
ciated with oral and topical retinoid treatments. In
order to determine if the antagonists in this study are
able to prevent the cutaneous toxicity produced by a
retinoid agonist in an in vivo model of topical irritation,
we tested compound 1 for its ability to inhibit the severe
cutaneous toxicity produced by the topical administra-
tion of TTNPB.12 As shown in Figure 1, topical appli-
cation of compound 1 alleviates the severe cutaneous
toxicity caused by TTNPB treatment in this clinically
relevant model. Thus, the compounds reported here
have the potential of being useful in the treatment of
the mucocutaneous toxicity that is almost universally
associated with both systemic and topical retinoid
treatment.
We also tested these compounds for their ability to
inhibit TTNPB-induced transactivation at each of the
RAR subtypes. Each compound was tested at varying
concentrations for its ability to inhibit the transcrip-
tional activation caused by TTNPB at a constant con-
centration of 10 nM as previously described.9b The half-
maximal inhibition concentration (IC50) values were
calculated and are listed in Table 1. Each of these com-
pounds acts as a full antagonist of TTNPB function at
all three RARs. Surprisingly, although these com-
pounds were somewhat RARb selective in binding, they
were most potent at RARg in inhibition of TTNPB
transactivation. This discrepancy may arise from the
fact that while binding assays are done using mono-
meric RARs, the transactivation assays are done in the
context of RAR±RXR heterodimers.
A possible therapeutic application of retinoid antago-
nists is to inhibit the skin irritation that is produced in
response to retinoid treatment. In particular, it would
In summary, we have described the synthesis and bio-
logical activity of a new series of RAR-speci®c antago-
nists, which are structurally less rigid than previously
described retinoid antagonists. These bind to the reti-
noic acid receptorsÐRARa, b, and gÐwith high a-
nity, and act as antagonists of TTNPB in receptor
transactivation assays. Moreover, in a hairless mouse
model of retinoid induced topical irritation, a repre-
sentative compound of this class is able to reverse the
topical irritation caused by topical administration of
TTNPB. Thus, these compounds may be useful as a
topical treatment for the mucocutaneous toxicity caused
by systemic retinoid medications such as 13-cis-retinoic
acid, as well as for treating other retinoid anatagonist
responsive diseases.
Table 1. RAR transcriptional activation, competitive binding and
antagonist activity for 2-phenyl-6,6-dimethyl-1-cyclohexene (1±3)
and 6-phenyl-3,3-dimethyl-1,5-cyclohexadiene (4 and 5) retinoid
antagonistsa,b,c
RAR trans. EC50 (nM)
RAR IC50 (nM)
Ecacy (%)
Compound number
RAR bind. Kd (nM)
a
b
g
a
b
g
ATRA
240
14
30
38
11
8
5
7
20
NA
7
NA
6
6
16
4
26
10
NDd ND ND
TTNPB
ND
ND
ND ND
ND ND
36
190121
435
135
NAe
17
190
NA
193109
31
(91)
210
(93)
220
(97)
350
(94)
129
(96)
ND
20
(97) (98)
22
(92) (96)
40
(96) (97)
27
(96) (95)
35
3
6
1
2
3
4
5
NA
148
NA
136
NA
875
NA
252
NA
>1000
NA
21
NA
36
NA
331
NA
31
NA
>1000
5
NA
4
3
NA
25
NA
53
NA
87
5
4
(94) (97)
ND ND
aSee refs 8 and 16.
bValues represent the mean of three determinations. Errors in this
assay are approximately 15% of the mean value.
cEcacy (%) is the percent transcriptional activity of 10 nM TTNPB
antagonized by each compound at the highest dose tested (10 mM).
dND, not determined.
Figure 1. Inhibition of systemic TTNPB-induced topical irritation of
hairless mouse skin by compound 1.12
eNA, not active (i.e., 0% ecacy at 104 nM).

768
R. L. Beard et al. / Bioorg. Med. Chem. Lett. 11 (2001) 765±768
8. Johnson, A. T.; Klein, E. S.; Gillett, S. J.; Wang, L.; Song,
Acknowledgements
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ligand binding data. We also thank Mr. Hai Nguyen for
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