Homologs of idoxifene: Variation of estrogen receptor binding and calmodulin antagonism with chain length

Article abstract of DOI:10.1021/jm9505472

A series of homologs of idoxifene [1a, (E)-1-[4-(N- pyrrolidinoethoxy)phenyl]-1-(4-iodophenyl)-2-phenyl-1-butene] and selected homologs of 4-iodotamoxifen [2a, (E)-1-[4-[(N-dimethylamino)ethoxy]phenyl]- 1-(4-iodophenyl)-2-phenyl-1-butene] with the side ch

Full text of DOI:10.1021/jm9505472

J . Med. Chem. 1996, 39, 999-1004
999
Notes
Hom ologs of Id oxifen e: Va r ia tion of Estr ogen Recep tor Bin d in g a n d
Ca lm od u lin An ta gon ism w ith Ch a in Len gth
Ian R. Hardcastle,* Martin G. Rowlands, Rachel M. Grimshaw, J ohn Houghton, and Michael J arman
The CRC Centre for Cancer Therapeutics at the Institute of Cancer Research, Cancer Research Campaign Laboratories,
Cotswold Road, Sutton, Surrey SM2 5NG, U.K.
Andrew Sharff and Stephen Neidle
CRC Biomolecular Structure Unit, The Institute of Cancer Research, Sutton, Surrey SM2 5NG, U.K.
Received J uly 24, 1995X
A series of homologs of idoxifene [1a , (E)-1-[4-(N-pyrrolidinoethoxy)phenyl]-1-(4-iodophenyl)-
2-phenyl-1-butene] and selected homologs of 4-iodotamoxifen [2a , (E)-1-[4-[(N-dimethylamino)-
ethoxy]phenyl]-1-(4-iodophenyl)-2-phenyl-1-butene] with the side chain (CH2)n varying in length
from n ) 3 (1b, 2b) to n ) 10 (1i, 2i) have been synthesized and tested for antagonism of the
calmodulin-dependent activity of cAMP phosphodiesterase and for binding affinity to rat uterine
estrogen receptor. Compared with 1a (IC50 ) 1.5 µM), the homologs showed a progressive
increase in calmodulin antagonism with a maximum inhibition at n ) 7-9 (1f-h ) (IC50 ) 0.2
µM), declining at n ) 10 (1i) to IC50 ) 1.6 µM. In the pyrrolidino series, estrogen receptor
binding affinity peaked at n ) 3 (1b, RBA ) 23; estradiol ) 100), declining by n ) 10 (1i) to
RBA ) 0.4, but the homolog n ) 8 (1g, RBA ) 3.5) was still comparable to tamoxifen (RBA )
3.9). A similar pattern of activity was seen for the dimethylamino counterparts. These
compounds represent a new class of antiestrogens with potent calmodulin antagonism.
In tr od u ction
We recently reported that the 3- and 4-carbon ho-
mologs of idoxifene [1a , (E)-1-[4-(N-pyrrolidinoethoxy)-
phenyl]-1-(4-iodophenyl)-2-phenyl-1-butene] and 4-io-
dotamoxifen [2a , (E)-1-[4-[(N-dimethylamino)ethoxy]-
phenyl]-1-(4-iodophenyl)-2-phenyl-1-butene] were more
potent calmodulin antagonists than the parent com-
pounds while retaining their estrogen receptor binding
affinity.1 We now report the extension of the homolo-
gous series and the identification of the 8-carbon ho-
mologs in both series as potent calmodulin antagonists.
Nonsteroidal antiestrogens such as 1a , in phase II
clinical trials,2 and its parent tamoxifen [3, trans-1-[4-
[2-(dim et h yla m in o)et h oxy]ph en yl]-1,2-diph en yl-1-
butene], widely used in the treatment of breast cancer,
are believed to act principally by the displacement of
the growth promoting hormone estradiol from its protein
receptor.3 Both compounds also exhibit a number of
hormone-independent effects which may contribute to
their therapeutic action. These include calmodulin
antagonism4-6 and inhibition of protein kinase C.7-10
The exact role of calmodulin in the functioning of the
estrogen receptor has not been defined. However, an
estrogen receptor-calmodulin-estradiol complex has
been identified, and its formation has been shown to be
antagonized by the addition of some antiestrogens.11
Also, a calmodulin-dependent protein tyrosine kinase
responsible for phosphorylating the estrogen receptor
has been identified.12 Antagonism of calmodulin-de-
pendent processes appears to correlate with cytotoxicity
in estrogen receptor (ER) positive cell lines for some
antiestrogens.1,6
The attractiveness of calmodulin as a target is en-
hanced by the fact that its structure is well known, and
so computerized molecular modeling can be used as part
of a rational drug design strategy. Our modeling studies
to date1,13 have used the open form of calmodulin seen
in X-ray crystal structures of the native protein.14-17
This work modeled the interactions of 3- and 4-carbon
homologs of 1a and 2a with calmodulin and suggested
that they would be more potent calmodulin antagonists.
The compounds 1b,c and 2b,c displayed increased an-
tagonism, although the maximum antagonism appeared
not to have been reached. More recent studies (to be
reported elsewhere) have taken the “closed” calmodulin
structures seen in crystal structures of complexes with
peptides and drugs,18-21 with analogous results to those
X Abstract published in Advance ACS Abstracts, J anuary 1, 1996.
0022-2623/96/1839-0999$12.00/0 © 1996 American Chemical Society

1000 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 4
Notes
Sch em e 1. Synthesis of Homologs of Idoxifene (1a ) and 4-Iodotamoxifen (2a )
with the open form.1,13 We have extended the homolo-
Ta ble 1. Antagonism of Calmodulin-Dependent cAMP
Phosphodiesterase and Receptor Binding Affinities for
Homologs of Idoxifene (1a )
gous series for 1a and 2a to explore further the
structure-activity relationships for calmodulin antago-
nism and estrogen receptor binding properties. In this
paper we report that the C7-9 homologs 1f-h and 2g
are potent calmodulin antagonists with estrogen recep-
tor binding affinities (RBA) similar to that of 3.
antagonism of CaM- potency relative binding
dependent PDE,a
IC50 ( SE (µM)
to parent
affinity
for ER
compound
n
compoundb
1a
1b
1c
1d
1e
1f
1g
1h
1i
2
3
4
5
6
7
8
9
10
1.5 ( 0.1
1.1 ( 0.1
1.0 ( 0.1
0.8 ( 0.08
0.5 ( 0.05
0.2 ( 0.05
0.2 ( 0.05
0.2 ( 0.05
1.6 ( 0.1
1
12
23
9
8
4
1.4
1.5
1.9
3.0
7.5
7.5
7.5
0.9
Resu lts a n d Discu ssion
Syn th esis. Hom ologs of Id oxifen e (1a ) a n d 4-Io-
d ota m oxifen (2a ). The synthesis of the 4-iodo ho-
mologs is outlined in Scheme 1. The procedure essen-
tially follows that reported for the synthesis of 3- and
4-carbon homologs 1b,c and 2b,c,1 substituting the
appropriate [(ω-haloalkyl)oxy]benzene 4d -i in the
Friedel-Crafts acylation of 2-phenylbutanoic acid to
give the 1,2-diarylbutanones 5d -i, respectively. Reac-
tion of the ketones 5d -i with 4-iodophenyllithium,
readily generated by treatment of 1,4-diiodobenzene
with 1 equiv of n-butyllithium, and subsequent dehy-
dration of the resulting tertiary alcohols gave the
triarylbutenes 6d -i, respectively, as a mixture of E and
Z isomers which were separable by fractional crystal-
lization. (Trans and cis are used in this paper to
designate the relative positions of the ethyl group and
the ring bearing the basic side chain.) The desired E
(trans) isomers were then treated with pyrrolidine or
dimethylamine to give the pyrrolidino compounds 1d -i
or dimethylamino compounds 2d -i, respectively.
Biologica l Eva lu a tion a n d Gen er a l Discu ssion .
Compounds were assayed for the inhibition of calmodu-
lin-dependent cyclic AMP phosphodiesterase (cAMP f
AMP) (none of the compounds inhibited the calmodulin-
independent component of cAMP phosphodiesterase
when assayed at the final concentrations of 10 and 20
µM), and the binding affinities of the compounds toward
rat uterine cytosolic estrogen receptor were determined
relative to estradiol (RBA ) 100) following the published
procedures.6,22 The results are summarized in Tables
1 and 2.
3
3.5
1.5
0.4
a
None of the compounds gave significant inhibition of the
calmodulin-independent activity of cAMP PDE when assayed at
b
final concentrations of 10 and 20 µM. Relative potency
)
IC50(parent)/IC50(compound).
Ta ble 2. Antagonism of Calmodulin-Dependent cAMP
Phosphodiesterase and Receptor Binding Affinities for
Homologs of 4-Iodotamoxifen (2a )
antagonism of CaM- potency relative binding
dependent PDE,a
IC50 ( SE (µM)
to parent
affinity
for ER
compound
n
compoundb
2a
2b
2c
2d
2e
2g
2i
2
3
4
5
6
2.3 ( 0.4
2.0 ( 0.2
2.2 ( 0.2
0.6 ( 0.05
0.4 ( 0.05
0.3 ( 0.05
1.2 ( 0.1
1
8
8
25
6
8
2
1.2
1.1
3.8
5.8
7.7
1.9
8
10
0.4
a
None of the compounds gave significant inhibition of the
calmodulin-independent activity of cAMP PDE when assayed at
b
final concentrations of 10 and 20 µM. Relative potency
)
IC50(parent)/IC50(compound).
amide antagonists 7 including the calmodulin antago-
nist W-7.23,24 In the case of the iodo-substituted com-
pounds, a jump in activity was seen from n ) 10 (IC50
) 4 µM) to n ) 12 (IC50 ) 0.7 µM), although no
explanation was offered.
In the pyrrolidino series there is a gradual rise in
relative potency from 1.4 to 3.0 for n ) 3-6 (1b-e)
followed by a sharp jump at n ) 7 (1f) to 7.5 (IC50 ) 0.2
µM). The 8- and 9-carbon homologs 1g,h display similar
activity. The calmodulin antagonism drops off sharply
at n ) 10 (1i) to that of the parent compound 1a .
Selected compounds in the dimethylamino series 2a -i
display a similar pattern with the C8 compound 2g being
the most potent (IC50 ) 0.30 µM). All the compounds
in this series displayed similar or worse activity com-
pared with their pyrrolidino counterparts.
Examination of the computer model of calmodulin
bound with 1a 1 does not readily explain why such a
dramatic increase in potency is seen from C6 to C7-9 or
why the activity should be much lower for C10. How-
ever, recent studies in which calmodulin is crystallized
with a peptide ligand, such as the calmodulin-binding
Elongation of the basic side chain has been seen to
produce an increase in potency in the naphthylsulfon-

Notes
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 4 1001
domain of smooth muscle myosin light chain kinase17,25
or protein kinase IIa,18 or the drug trifluoperazine20,21
have shown calmodulin in a more compact globular
structure with the ligand enclosed by the N- and
C-terminal lobes. It seems likely that the binding of
1a or 2a and their homologs to calmodulin would
produce a similar conformational change in the protein.
Molecular modeling and crystallographic studies are
underway in our laboratories to provide detailed struc-
tural information which can be used to rationalize these
interactions.
Estrogen receptor binding affinity for both series
drops with extending chain length beyond C6. However,
the most potent calmodulin antagonists (1f-h and 2g)
still have RBA values similar to that of 3. This suggests
that these compounds may be capable of antagonizing
both estrogen- and calmodulin-dependent processes in
vivo. Receptor binding affinity is lost at C10 (1i and 2i)
in both series.
1-Ch lor o-8-p h en oxyocta n e (4g): 9.20 g, 72%; bp 180 °C
(0.3 mmHg); 1H NMR (CDCl3) δ 1.28-1.85 (m, 12, CH2(CH2)6-
CH2), 3.54 (t, J ) 6.4 Hz, 2, ClCH2), 3.94 (t, J ) 6.4 Hz, 2,
OCH2), 6.86-6.92 (m, 3, ArH), 7.20-7.31 (m, 2, ArH); MS (EI)
m/ z 240 (M+, 35).
1-Ch lor o-9-p h en oxyn on a n e (4h ): 4.6 g, 61%; bp 175 °C
1
(0.4 mmHg); H NMR (CDCl3) δ 1.26-1.49 (10, m, O(CH2)2-
(CH2)5(CH2)2Cl), 1.71-1.83 (m, 4, OCH2CH2(CH2)5CH2CH2Cl),
3.53 (t, J ) 6.4 Hz, 2, ClCH2), 3.95 (2, t, J ) 6.4 Hz, OCH2),
6.88-6.96 (m, 3, ArH), 7.24-7.31 (m, 2, ArH); MS (EI) m/ z
254 (M+ - 1, 50).
1-Ch lor o-10-p h en oxyd eca n e (4i): 9.8 g, 69%; bp 185 °C
(0.2 mmHg); 1H NMR (CDCl3) δ 1.2-1.8 (m, 16, OCH2(CH2)8-
CH2Cl), 3.51 (t, J ) 6.4 Hz, 2, ClCH2), 3.93 (t, J ) 6.4 Hz, 2,
OCH2), 6.82-6.94 (m, 3, ArH), 7.20-7.30 (m, 2, ArH); MS (EI)
m/ z 268 (M+, 5).
1-Br om o-7-p h en oxyh ep ta n e (4f). A mixture of phenol
(1.7 g, 10 mmol), 1,7-dibromoheptane (10 g, 39 mmol), NaOH
solution (1 M, 20 mL), and tetrabutylammonium hydrogen
sulfate (0.05 g) was refluxed for 16 h and then diluted with
ether (50 mL) and partitioned. The ether layer was washed
with HCl (1 M, 25 mL) and water (25 mL), dried (Na2SO4),
and concentrated. Distillation gave 4f as a colorless oil (2.38
g, 90%): bp 180 °C (0.2 mmHg); 1H NMR (CDCl3) δ 1.21-
1.54 and 1.73 -1.93 (m, 10, CH2(CH2)5CH2), 3.42 (t, J ) 6.4
Hz, 2, CH2Br), 3.95 (t, J ) 6.4 Hz, 2, PhOCH2), 6.87-6.96 (m,
2, ArH), 7.24-7.32 (m, 3, ArH); MS (EI) m/ z 270 (M+ - 1,
20).
Gen er a l P r ep a r a tion of 1-[4-(ω-Ch lor oa lk oxy)p h en yl]-
2-p h en yl-1-bu ta n on es 5d-i. To a stirred solution of 2-phen-
ylbutyric acid (20 mmol) and trifluoroacetic anhydride (5 mL)
was added (ω-haloalkoxy)benzene 4d -i (24 mmol). The
resulting mixture was stirred for 16 h and then poured into
saturated aqueous NaHCO3 solution (30 mL) and extracted
with ether (30 mL). The organic extracts were washed with
water (2 × 30 mL), dried (MgSO4), and concentrated.
1-[4-[(5-Ch lor op e n t yl)oxy]p h e n yl]-2-p h e n yl-1-b u t a -
n on e (5d ). Crystallization (hexane) gave 5d as fine white
needles (5.49 g, 80%): mp 60-61 °C; 1H NMR (CDCl3) δ 0.87
(t, J ) 7.5 Hz, 3, CH2CH3), 1.56-1.90 (m, 7, CH2(CH2)3CH2,
part of CHOCH2), 2.11-2.22 (m, 1, part of CHOCH2), 3.54 (t,
J ) 6.4 Hz, 2, ClCH2), 3.96 (t, J ) 6.4 Hz, 2, OCH2), 4.38 (t, J
) 7.3 Hz, 1, CHOCH2), 6.82 (d, J ) 9.0 Hz, 2, ArH ortho to
OCH2), 7.13-7.28 (m, 5, PhH), 7.93 (d, J ) 8.8 Hz, 2, ArH
meta to OCH2). Anal. (C21H25O2Cl) C, H, Cl.
1-[4-[(6-Ch lor oh e xyl)oxy]p h e n yl]-2-p h e n yl-1-b u t a -
n on e (5e). Chromatography (CH2Cl2-hexane, 1:5) gave 5e
as white amorphous crystals (6.00 g, 84%): mp 43-45 °C
(hexane); 1H NMR (CDCl3) δ 0.87 (t, J ) 7.4 Hz, 3, CH2CH3),
1.45-1.87 (m, 9, CH2(CH2)4CH2, part of CHOCH2), 2.11-2.22
(m, 1, part of CHOCH2), 3.52 (t, J ) 7.7 Hz, 2, ClCH2), 3.95 (t,
J ) 6.4 Hz, 2, OCH2), 4.38 (t, J ) 7.3 Hz, 1, CHOCH2), 6.82
(d, J ) 7.9 Hz, 2, ArH ortho to OCH2), 7.16-7.28 (m, 5, PhH),
7.93 (d, J ) 9.0 Hz, 2, ArH meta to OCH2); MS (CI) m/ z 359
(M+, 12). Anal. (C22H27O2Cl) C, H, Cl.
1-[4-[(10-Ch lor od e cyl)oxy]p h e n yl]-2-p h e n yl-1-b u t a -
n on e (5i). Chromatography (CH2Cl2-petroleum ether, 1:5)
gave 5i as white crystals (5.83 g, 70%): mp 42-44 °C (hexane);
1H NMR (CDCl3) δ 0.89 (t, J ) 7.3 Hz, 3, CH2CH3), 1.20-1.92
(m, 17, CH2(CH2)8CH2, part of CHOCH2), 2.13-2.27 (m, 1, part
of CHOCH2), 3.53 (t, J ) 6.6 Hz, 2, ClCH2), 3.96 (t, J ) 6.5
Hz, 2, OCH2), 4.39 (t, J ) 7.3 Hz, 1, CHOCH2), 6.84 (d, J )
8.8 Hz, 2, ArH ortho to OCH2), 7.18-7.32 (m, 5, PhH), 7.92
(d, J ) 8.9 Hz, 2, ArH meta to OCH2); MS (CI) m/ z 415 (M+,
72). Anal. (C26H35O2Cl) C, H, Cl.
1-[4-[(8-Ch lor ooct yl)oxy]p h e n yl]-2-p h e n yl-1-b u t a -
n on e (5g). 2-Phenylbutyric acid (7.48 g, 46 mmol), 4g (9.1 g,
38 mmol), and trifluoroacetic anhydride (7.5 mL, 52 mmol)
were treated as in the general procedure. Chromatography
(CH2Cl2-petroleum ether, 1:5) gave 5g as white crystals (11.86
g, 81%): mp 56-57 °C (hexane); 1H NMR (CDCl3) δ 0.87 (t, J
) 7.3 Hz, 3, CH2CH3), 1.26-1.50 and 1.69-1.90 (m, 14,
CH2(CH2)6CH2Cl, part of CH2CH3), 2.08-2.26 (m, 1, part of
CH2CH3), 3.51 (t, J ) 6.6 Hz, 2, (CH2)6CH2Cl), 3.94 (t, J ) 6.6
Hz, 2, OCH2(CH2)6), 4.38 (t, J ) 7.3 Hz, 1, CHCO), 6.83 (d, J
) 8.8 Hz, 2, ArH meta to OCH2), 7.15-7.20 (m, 1, PhH), 7.20-
Con clu sion s
Extension of the homologous series for 1a and 2a has
revealed that elongation of the basic side chain results
in greater antagonistic potency for calmodulin while the
estrogen receptor binding affinity falls. The C7-9 ho-
mologs in both series (1f-h and 2g) are potent calmod-
ulin antagonists (IC50 ) 0.2 µM) and have similar
estrogen receptor binding affinities to 3 (RBA ∼ 2).
These compounds represent a new class of antiestrogens
which are potent calmodulin antagonists. They will be
useful in elucidating the complex interactions of calmod-
ulin and the estrogen receptor and may be novel
therapeutic agents for the treatment of cancer.
Exp er im en ta l Section
Ch em ica l Meth od s. Gen er a l P r oced u r es. 1H NMR
spectra (internal Me4Si) were obtained with a Bruker AC250
instrument. Melting points were obtained on a Reichert
hotstage and are uncorrected. Chromatography refers to flash
column chromatography on silica gel (Merck 15111) with the
eluant indicated applied at a positive pressure of 0.5 atm. All
reactions performed under an inert atmosphere were carried
out in oven-dried glassware (110 °C, 24 h). Ether refers to
diethyl ether. Hexane for chromatography was purchased
from Romil (super purity grade). Anhydrous tetrahydrofuran
(THF) was obtained by distillation from potassium and ben-
zophenone. Purification of E and Z geometrical isomers was
monitored by 1H NMR spectroscopy and was carried out until
none of the undesired isomer could be detected. Elemental
analyses were determined by CHN Analysis Ltd., South
Wigston, Leicester, England.
Gen er a l Meth od for P r ep a r a tion of 1-Ch lor o-ω-p h e-
n oxya lk a n es. A two-phase mixture of phenol (5 g, 53 mmol),
R,ω-dihaloalkane (30 mL), tetrabutylammonium hydrogen
sulfate (0.3 g, 1 mmol), and 3 M NaOH (25 mL) was heated to
reflux for 16 h. The mixture was diluted with ether (50 mL)
and washed with HCl (1 M, 30 mL) and water (2 × 25 mL).
The ether layer was dried (MgSO4) and concentrated. Chro-
matography (CH2Cl2-hexane, 1:10) gave the products as
colorless oils.
1-Ch lor o-5-p h en oxyp en ta n e (4d ): 9.32 g, 94%; bp 110
°C (0.1 mmHg); 1H NMR (CDCl3) δ 1.55-1.91 (m, 6, CH2(CH2)3-
CH2), 3.56 (t, J ) 6.7 Hz, 2, ClCH2), 3.95 (t, J ) 6.3 Hz, 2,
OCH2), 6.87-6.96 (m, 3, ArH), 7.24-7.31 (m, 2, ArH); MS (EI)
m/ z 198 (M+, 65).
1-Ch lor o-6-p h en oxyh exa n e (4e): 9.21 g, 82%; bp 130 °C
1
(0.1 mmHg); H NMR (CDCl3) δ 1.43-1.83 (m, 8, CH2(CH2)4-
CH2), 3.54 (t, J ) 6.7 Hz, 2, ClCH2), 3.95 (t, J ) 6.4 Hz, 2,
OCH2), 6.85-6.96 (m, 3, ArH), 7.22-7.31 (m, 2, ArH); MS (EI)
m/ z 212 (M+, 15).

1002 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 4
Notes
7.28 (m, 4, PhH), 7.93 (d, J ) 8.8 Hz, 2, ArH ortho to OCH2);
IR (liquid, cm-1) 2933, 2858, 1673, 1600, 1574, 1510; MS (CI)
m/ z 387 (M+ + 1, 10). Anal. (C24H31O2Cl) C, H, Cl.
MS (EI) m/ z 604 (M+, 10); accurate mass (C29H32O79BrI) found
602.0681, calcd 602.0686.
(E)-1-[4-[(8-Ch lor ooctyl)oxy]p h en yl]-1-(4-iod op h en yl)-
2-p h en yl-1-bu ten e (6g). 1,4-Diiodobenzene (9.9 g, 30 mmol)
in THF (100 mL), n-butyllithium (1.6 M in hexane, 18.75 mL,
30 mmol), and 5g (11.18 g, 29 mmol) in tetrahydrofuran (50
mL) were treated as in the general procedure. Chromatogra-
phy (CH2Cl2-hexane, 1:10) followed by fractional crystalliza-
tion gave 6g as white crystals (5.35 g, 31%): mp 55-58 °C;
1H NMR (CDCl3) δ 0.89 (t, J ) 7.5 Hz, 3, CH2CH3), 1.26-1.46
and 1.60-1.80 (m, 12, CH2(CH2)6CH2), 2.415 (q, J ) 7.4 Hz,
2, CH2CH3), 3.50 (t, J ) 6.6 Hz, 2, CH2Cl), 3.79 (t, J ) 6.6 Hz,
2, OCH2), 6.505 (d, J ) 8.7 Hz, 2, ArH meta to OCH2), 6.705
(d, J ) 8.7 Hz, 2, ArH meta to OCH2), 6.965 (d, J ) 8.4 Hz,2,
7.07-7.19 (m, 5, PhH), 7.65 (d, J ) 8.2 Hz, ArH ortho to I);
υmax (film) 2931, 2857, 1606, 1509; MS (CI) m/ z 574 (M+ + 1,
10). Anal. (C30H34OClI) C, H, Cl, I.
1-[4-[(9-Ch lor on on yl)oxy]p h e n yl]-2-p h e n yl-1-b u t a -
n on e (5h ). 2-Phenylbutyric acid (3.28 g, 20 mmol), 4h (4.60
g, 18 mmol), and trifluoroacetic anhydride (20 mL) were
treated as above. Chromatography (CH2Cl2-petroleum ether,
3:10) gave 5h as white crystals (5.83 g, 70%): mp 42-44 °C
(hexane); 1H NMR (CDCl3) δ 0.89 (t, J ) 7.4 Hz, 3, CH2CH3),
1.22-1.50 and 1.71-1.89 (m, 15, CH2(CH2)7CH2Cl, part of CH2-
CH3), 2.13-2.24 (m, 1, part of CH2CH3), 3.53 (t, J ) 6.8 Hz,
CH2Cl), 3.96 (t, J ) 6.5 Hz, OCH2), 4.40 (t, J ) 7.3 Hz, 1,
CHCO), 6.84 (d, J ) 8.8 Hz, 2, ArH meta to OCH2), 7.17-7.32
(m, 5, Ph), 7.95 (d, J ) 9.0 Hz, 2, ArH ortho to OCH2); MS
(CI) m/ z 401 (M+, 15). Anal. (C25H33O2Cl) C, H, Cl.
1-[4-[(7-Br om oh e p t yl)oxy]p h e n yl]-2-p h e n yl-1-b u t a -
n on e (5f). 2-Phenylbutyric acid (2.02 g, 10 mmol), 4f (1.70 g,
6.3 mmol), and trifluoroacetic anhydride (10 mL) were treated
as above. Chromatography (CH2Cl2-petroleum ether, 1:4)
gave 5f as a colorless oil (2.85 g, 77%): mp 41-43 °C (hexane);
1H NMR (CDCl3) δ 0.89 (m, 3, CH2CH3), 1.25-1.51 and 1.69-
1.92 (m, 10, OCH2(CH2)5CH2Br), 2.03-2.27 (m, 2, CH2CH3),
3.41 (t, J ) 6.8 Hz, 2, CH2Br), 3.96 (t, J ) 6.5 Hz, OCH2), 4.40
(t, J ) 7.2 Hz, 1, COCH), 6.84 (d, J ) 8.9 Hz, 2, ArH ortho to
OCH2), 7.15-7.35 (m, 5, PhH), 7.95 (d, J ) 8.8 Hz, 2, ArH
meta to OCH2); MS (EI) m/ z 418 (M+, 100); accurate mass
(C23H29O2Br) found 417.1423, calcd 417.1419.
Gen er a l P r ep a r a t ion of (E)-1-[4-(ω-Ch lor oa lk oxy)-
p h en yl]-1-(4-iod op h en yl)-2-p h en yl-1-bu ten es 6d -i. To a
solution of 1,4-diiodobenzene (3.629 g, 11 mmol) in THF (30
mL) was added n-butyllithium (1.6 M in hexane, 6.9 mL, 11
mmol) at -78 °C. The resulting mixture was stirred for 10
min; then a solution of the ketone 5d -i (11 mmol) in THF (20
mL) was added, and stirring continued at -78 °C for 1 h
followed by 16 h at ambient temperature. Ammonium chloride
solution (2 mL) was added; the mixture was diluted with ether
(100 mL) and then washed with brine (100 mL) and water (2
× 100 mL). The organic layer was dried (MgSO4) and
concentrated. The residues were dissolved in ethanol (30 mL)
and HCl (30%, 10 mL). The mixture was heated to reflux for
90 min, allowed to cool, diluted with ether (50 mL), and washed
with water (50 mL), aqueous sodium thiosulfate solution (5
M, 50 mL), and water (50 mL). The organic layer was dried
(MgSO4) and concentrated.
(E)-1-[4-[(5-Ch lor open tyl)oxy]ph en yl]-1-(4-iodoph en yl)-
2-p h en yl-1-bu ten e (6d ). Chromatography (CH2Cl2-hexane,
1:5) followed by fractional crystallization (ethanol) gave 6d as
white crystals (2.402 g, 47%): mp 94-96 °C; 1H NMR (CDCl3)
δ 0.89 (t, J ) 7.5 Hz, 3, CH2CH3), 1.50-1.82 (m, 6, CH2(CH2)3-
CH2), 2.42 (q, J ) 7.4 Hz, 2, CH2CH3), 3.52 (t, J ) 6.6 Hz, 2,
CH2Cl), 3.81 (t, J ) 6.2 Hz, 2, CH2O), 6.51 (d, J ) 8.7 Hz, 2,
ArH ortho to OCH2), 6.71 (d, J ) 8.5 Hz, 2, ArH meta to OCH2),
6.97 (d, J ) 8.4 Hz, 2, ArH meta to I), 7.07-7.19 (m, 5, PhH),
7.65 (d, J ) 8.4 Hz, 2, ArH ortho to I). Anal. (C27H28OClI) C,
H, Cl, I.
(E)-1-[4-[(9-Ch lor on on yl)oxy]p h en yl]-1-(4-iod op h en yl)-
2-p h en yl-1-bu ten e (6h ). Chromatography (CH2Cl2-hexane,
1:5) followed by fractional crystallization (hexane) gave 6h as
1
white crystals (1.96 g, 30%): mp 73-75 °C; H NMR (CDCl3)
δ 0.91 (t, J ) 7.5 Hz, 3, CH2CH3), 1.25-1.88 (m, 14, CH2(CH2)7-
CH2), 2.43 (q, J ) 7.4 Hz, 2, CH2CH3), 3.52 (t, J ) 6.7 Hz, 2,
CH2Cl), 3.81 (t, J ) 6.6 Hz, 2, CH2O), 6.53 (d, J ) 8.8 Hz, 2,
ArH ortho to OCH2
), 6.73 (d, J ) 8.8 Hz, 2, ArH meta to OCH2),
6.98 (d, J ) 8.3 Hz, 2, ArH meta to I), 7.09-7.21 (m, 5, PhH),
7.67 (d, J ) 8.5 Hz, 2, ArH ortho to I); MS (EI) m/ z 586 (M+
- 1, 100). Anal. (C31H36OClI) H, Cl, I; C: calcd, 63.43; found,
64.11.
(E)-1-[4-[(10-Ch lor odecyl)oxy]ph en yl]-1-(4-iodoph en yl)-
2-p h en yl-1-bu ten e (6i). Chromatography (20% CH2Cl2, hex-
ane) followed by fractional crystallization (ethanol) gave 6i as
white crystals (2.35 g, 35%): mp 75-78 °C; 1H NMR (CDCl3)
δ 0.89 (t, J ) 7.5 Hz, 3, CH2CH3), 1.26-1.77 (m, 16, CH2(CH2)8-
CH2), 2.42 (q, J ) 7.4 Hz, 2, CH2CH3), 3.51 (t, J ) 6.8 Hz, 2,
CH2Cl), 3.79 (t, J ) 6.6 Hz, 2, CH2O), 6.51 (d, J ) 8.5 Hz, 2,
ArH ortho to OCH2), 6.70 (d, J ) 8.7 Hz, 2, ArH meta to OCH2),
6.97 (d, J ) 8.4 Hz, 2, ArH meta to I), 7.07-7.15 (m, 5, PhH),
7.65 (d, J ) 8.5 Hz, 2, ArH ortho to I). Anal. (C32H38OClI) C,
H, Cl, I.
Gen er a l P r oced u r e for P r ep a r a tion of (E)-1-[4-(ω-N-
P yr r olid in oa lk oxy)p h en yl]-1-(4-iod op h en yl)-2-p h en yl-1-
bu ten es 1d -i. A mixture of butene 6d -i (1 mmol), pyrroli-
dine (2 mL), and ethanol (10 mL) was heated in a bomb at
100 °C for 4 h and then concentrated. Chromatography (ether)
gave the title compounds.
(E)-1-[4-[(5-N-P yr r olid in op en tyl)oxy]p h en yl]-1-(4-iod o-
p h en yl)-2-p h en yl-1-bu ten e (1d ): white crystals (0.60 g,
98%); mp 88-91 °C; 1H NMR (CDCl3) δ 0.89 (t, J ) 7.5 Hz, 3,
CH2CH3), 1.18-1.80 and 2.37-2.57 (m, 16, CH2(CH2)3CH2N,
N(CH2CH2)2), 3.79 (t, J ) 6.5 Hz, 2, OCH2), 6.50 (d, J ) 8.7
Hz, 2, ArH ortho to OCH2), 6.70 (d, J ) 8.7 Hz, 2, ArH meta
to OCH2), 6.97 (d, J ) 8.3 Hz, 2, ArH meta to I), 7.07-7.19
(m, 5, PhH), 7.65 (d, J ) 8.2 Hz, 2, ArH ortho to I); MS m/ z
565 (M+, 35). Anal. (C31H36NOI) C, H, N, I.
(E)-1-[4-[(6-N-P yr r olid in oh exyl)oxy]p h en yl)-1-(4-iod o-
p h en yl)-2-p h en yl-1-bu ten e (1e): off-white crystals (0.56 g,
96%); mp 38-42 °C; 1H NMR (CDCl3) δ 0.91 (t, J ) 7.4 Hz, 3,
CH2CH3), 1.63-1.79 and 2.37-2.50 (m, 20, N(CH2CH2)2, CH2-
CH3, CH2(CH2)4CH2N), 3.81 (t, J ) 6.5 Hz, 2, OCH2), 6.52 (d,
J ) 8.8 Hz, 2, ArH ortho to OCH2), 6.72 (d, J ) 8.8 Hz, 2, ArH
meta to OCH2), 6.98 (d, J ) 8.5 Hz, 2, ArH meta to I), 7.08-
7.16 (m, 5, PhH), 7.66 (d, J ) 8.3 Hz, 2, ArH ortho to I); MS
m/ z 579 (M+, 80). Anal. (C32H38NOI) C, H, N; I: calcd, 22.93;
found, 22.11.
(E)-1-[4-[(7-N-P yr r olid in oh ep tyl)oxy]p h en yl]-1-(4-iod o-
p h en yl)-2-p h en yl-1-bu ten e (1f): white crystals (0.136 g,
91%); mp 45-50 °C; 1H NMR (CDCl3) δ 0.85-0.95, 1.26-1.88
and 2.36-2.47 (m, 25, N(CH2CH2)2, CH2CH3, CH2(CH2)5CH2N),
3.80 (t, J ) 6.5 Hz, 2, OCH2), 6.52 (d, J ) 8.8 Hz, 2, ArH ortho
to OCH2), 6.72 (d, J ) 8.7 Hz, 2, ArH meta to OCH2), 6.98 (d,
J ) 8.2 Hz, 2, ArH meta to I), 7.11-7.27 (m, 5, PhH), 7.66 (d,
J ) 8.3 Hz, 2, ArH ortho to I); MS (FAB) m/ z 594 (M+, 100);
accurate mass (C33H42NOI) found 595.2816, calcd 595.2811.
(E)-1-[4-[(8-N-P yr r olid in ooctyl)oxy]p h en yl]-1-(4-iod o-
p h en yl)-2-p h en yl-1-bu ten e (1g). A mixture of butene 6g
(E)-1-[4-[(6-Ch lor oh exyl)oxy]p h en yl]-1-(4-iod op h en yl)-
2-p h en yl-1-bu ten e (6e). Chromatography (CH2Cl2-hexane,
1:5) followed by fractional crystallization (ethanol) gave 6e as
white crystals (2.903 g, 53%): mp 58-62 °C; 1H NMR (CDCl3)
δ 0.89 (t, J ) 7.5 Hz, 3, CH2CH3), 1.36-1.80 (m, 8, CH2(CH2)4-
CH2), 2.41 (q, J ) 7.5 Hz, 2, CH2CH3), 3.51 (t, J ) 6.4 Hz, 2,
CH2Cl), 3.80 (t, J ) 6.4 Hz, 2, CH2O), 6.51 (d, J ) 9.0 Hz, 2,
ArH ortho to OCH2), 6.71 (d, J ) 9.0 Hz, 2, ArH meta to OCH2),
6.97 (d, J ) 8.5 Hz, 2, ArH meta to I), 7.06-7.20 (m, 5, PhH),
7.64 (d, J ) 8.5 Hz, 2, ArH ortho to I); MS (FAB) m/ z 545
(M+, 100). Anal. (C28H30OClI) C, H, N, Cl, I.
(E)-1-[4-[(7-Br om oh eptyl)oxy]ph en yl]-1-(4-iodoph en yl)-
2-p h en yl-1-bu ten e (6f). Chromatography (CH2Cl2-hexane,
1:10) followed by fractional crystallization (ethanol) gave 6f
as white crystals (0.197 g, 9%): mp 54-57 °C; 1H NMR (CDCl3)
δ 0.91 (t, J ) 7.3 Hz, 3, CH2CH3), 1.30-1.51 and 1.62-1.97
(m, 10, CH2(CH2)5CH2), 2.44 (q, J ) 7.3 Hz, 2, CH2CH3), 3.40
(t, J ) 8.6 Hz, 2, CH2Br), 3.81 (t, J ) 8.6 Hz, 2, OCH2), 6.53
(d, J ) 8.8 Hz, 2, ArH meta to OCH2), 6.73 (d, J ) 9 Hz, 2,
ArH meta to OCH2), 6.99 (d, J ) 8.5 Hz, 2, ArH meta to I),
7.12-7.25 (m, 5, PhH), 7.67 (d, J ) 8.5 Hz, 2, ArH ortho to I);

Notes
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 4 1003
(2.0 g, 3.5 mmol), pyrrolidine (15 mL), and ethanol (75 mL)
was heated in a bomb at 100 °C for 4 h and then concentrated.
Chromatography (ether) gave 1g as a slightly brown oil (1.92
g, 90%); recrystallization (methanol) gave the title compound
J ) 8.1 Hz, 2, ArH ortho to I); MS m/ z 610 (M+, 100). Anal.
(C34H44NOI) C, H, N, I.
Estr ogen Recep tor Bin d in g Assa y. The affinity of the
antiestrogens for the ER was measured using a competitive
binding assay as described by Wakeling.22 Immature rat
cytosol was incubated at 4 °C for 16 h with 5 nM 17â-[2,4,6,7-
3H]estradiol in the presence of increasing amounts (0.1-
100 000 nM) of test compounds dissolved in dimethylforma-
mide or unlabeled estradiol (control). The nonspecific binding
was quantified by a parallel set of tubes containing a 200-fold
excess (with respect to [3H]estradiol) of diethylstilboestrol.
Unbound compounds were removed with dextran-coated char-
coal, and the receptor-bound [3H]estradiol was determined. The
relative concentrations of estradiol and test compound required
to achieve 50% inhibition of [3H]estradiol binding give the RBA
which is [IC50 (estradiol)/IC50 (test compound)] × 100.
Ca lm od u lin An ta gon ism . This was determined using the
calmodulin-dependent cyclic AMP phosphodiesterase as previ-
ously described.6,26 The enzyme was assayed using 8-[3H]-
cAMP as substrate. The tritiated AMP formed during the
incubation was converted into tritiated adenosine by the 5′-
nucleotidase in snake venom. Product nucleosides were
separated from unreacted substrate by batch elution with
Dowex anion exchange resin with 3 mM acetic acid. The basal
activity of cAMP phosphodiesterase (calmodulin independent)
was determined by adding 1 mM EGTA to the assay medium.
Assays were carried out in the presence and absence of
different concentrations of the compounds dissolved in di-
methyl sulfoxide. The results are expressed as the concentra-
tion of inhibitor giving 50% inhibition of the calmodulin-
dependent cAMP phosphodiesterase and are the mean of
triplicate determinations ( standard error.
1
as off-white crystals: mp 55-58 °C; H NMR (CDCl3) δ 0.89
(t, J ) 7.4 Hz, 3, CH2CH3), 1.22-1.80 and 1.60-1.80 (m, 16,
CH2(CH2)6CH2, N(CH2CH2)2), 2.31-2.50 (m, 8, N(CH2CH2)2,
CH2CH3, NCH2), 3.78 (t, J ) 6.5 Hz, 2, OCH2), 6.51 (d, J )
8.7 Hz, 2, ArH ortho to OCH2), 6.705 (d, J ) 8.6 Hz, ArH meta
to OCH2), 6.965 (d, J ) 8.2 Hz, ArH meta to I), 7.07-7.19 (m,
5, PhH), 7.645 (d, J ) 8.2 Hz, ArH ortho to I); m/ z (EI) 608
(M+, 20). Anal. (C34H42NOI) C, H, N, I.
(E)-1-[4-[(9-N-P yr r olid in on on yl)oxy]p h en yl]-1-(4-iod o-
p h en yl)-2-p h en yl-1-bu ten e (1h ): off-white crystals (0.588 g,
96%); mp 69-71 °C; 1H NMR (CDCl3) δ 0.91 (t, J ) 7.4 Hz, 3,
CH2CH3), 1.28-1.80 (m, 18, N(CH2CH2)2, CH2(CH2)7CH2),
2.37-2.48 (m, 8, N(CH2CH2)2, CH2CH3, CH2N), 3.81(t, J ) 6.5
Hz, 2, OCH2), 6.53 (d, J ) 8.7 Hz, 2, ArH ortho to OCH2), 6.73
(d, J ) 8.8 Hz, 2, ArH meta to OCH2), 6.98 (d, J ) 8.5 Hz, 2,
ArH meta to I), 7.08-7.15 (m, 5, PhH), 7.66 (d, J ) 8.4 Hz, 2,
ArH ortho to I); MS m/ z 621 (M+, 20). Anal. (C35H44NOI) C,
H, N, I.
(E)-1-[4-[(10-N-P yr r olid in od ecyl)oxy]p h en yl]-1-(4-iod o-
p h en yl)-2-p h en yl-1-bu ten e (1i): white needles (0.579 g,
1
91%); mp 75-76 °C (MeOH); H NMR (CDCl3) δ 0.90 (t, J )
7.3 Hz, 3, CH2CH3), 1.21-1.81 (m, 20, N(CH2CH2)2, CH2(CH2)8-
CH2), 2.40-2.52 (m, 8, N(CH2CH2)2, CH2CH3, CH2N), 3.80 (t,
J ) 6.5 Hz, 2, OCH2), 6.53 (d, J ) 8.2 Hz, 2, ArH ortho to
OCH2), 6.72 (d, J ) 8.5 Hz, 2, ArH meta to OCH2), 6.98 (d, J
) 7.7 Hz, 2, ArH meta to I), 7.08-7.17 (m, 5, PhH), 7.66 (d, J
) 7.6 Hz, 2, ArH ortho to I); MS m/ z 636 (M+, 100). Anal.
(C36H46NOI) C, H, N, I.
Gen er a l P r oced u r e for P r ep a r a tion of (E)-1-[4-[ω-(N-
Dim eth ylam in o)alkoxy]ph en yl]-1-(4-iodoph en yl)-2-ph en -
yl-1-bu ten es 2d -i. A mixture of butene 6d -i (1 mmol) and
dimethylamine (30% in ethanol, 20 mL) was heated in a bomb
at 100 °C for 4 h and then concentrated. Chromatography
(methanol-ether, 1:20) gave 2d -i.
Ack n ow led gm en t. This work was funded by grants
from the Cancer Research Campaign and the Medical
Research Council. Mass spectrometry was provided by
the ULICS at the London School of Pharmacy.
(E)-1-[4-[[5-(N-Dim et h yla m in o)p en t yl]oxy]p h en yl]-1-
(4-iod op h en yl)-2-p h en yl-1-bu ten e (2d ): colorless oil (0.412
g, 76%); H NMR (CDCl3) δ 0.89 (t, J ) 7.3 Hz, 3, CH2CH3),
Refer en ces
1
(1) Hardcastle, I. R.; Rowlands, M. G.; Houghton, J .; Parr, I. B.;
Potter, G. A.; J arman, M.; Edwards, K. J .; Laughton, C. A.;
Trent, J . O.; Neidle, S. Rationally designed analogues of tamox-
ifen with improved calmodulin antagonism. J . Med. Chem. 1995,
38, 241-248.
(2) Haynes, B. P.; Quigley, M.; Doody, D. A.; Clarkson, A.; Griggs,
L. J .; Dowsett, M.; J arman, M. Pharmacokinetics and pharma-
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1.40-1.86 (m, 8, N(CH2)4CH2), 2.23 (s, 6, (CH3)2N), 2.42 (q, J
) 6.5 Hz, 2, CH2CH3), 3.80 (t, J ) 6.4 Hz, OCH2), 6.50 (d, J )
8.9 Hz, 2, ArH ortho to OCH2), 6.70 (d, J ) 8.8 Hz, 2, ArH
meta to OCH2), 6.96 (d, J ) 8.2 Hz, 2, ArH meta to I), 7.06-
7.19 (m, 5, PhH), 7.64 (d, J ) 8.3 Hz, 2, ArH ortho to I); MS
m/ z 539 (M+, 100). Anal. (C29H34NOI) H, N, I; C: calcd, 64.56;
found, 64.11.
(E)-1-[4-[[6-(N-Dim eth yla m in o)h exyl]oxy]p h en yl]-1-(4-
iod op h en yl)-2-p h en yl-1-bu ten e (2e): colorless oil (0.515 g,
93%); 1H NMR (CDCl3) δ 0.91 (t, J ) 7.4 Hz, 3, CH2CH3), 1.15-
1.52 and 1.68 (m, brs, 10, N(CH2)5CH2), 2.21 (s, 6, N(CH3)2),
3.81 (t, J ) 6.4 Hz, 2, OCH2), 6.53 (d, J ) 8.8 Hz, 2, ArH ortho
to OCH2), 6.72 (d, J ) 8.8 Hz, 2, ArH meta to OCH2), 6.98 (d,
J ) 8.5 Hz, 2, ArH meta to I), 7.09-7.21 (m, 5, PhH), 7.67 (d,
J ) 8.3 Hz, 2, ArH ortho to I); MS m/ z 553 (M+, 100). Anal.
(C30H36NOI) C, H, N; I: calcd, 22.93; found 22.11.
(E)-1-[4-[[8-(N-Dim eth yla m in o)octyl]oxy]p h en yl]-1-(4-
iod op h en yl)-2-p h en yl-1-bu ten e (2g). 6g (0.51 g, 0.89 mmol)
gave 2g as off-white crystals (0.446 g, 86%): mp 65-68 °C;
1H NMR (CDCl3) δ 0.91 (t, J ) 7.4 Hz, 3, CH2CH3), 1.21-1.83
(m, 12, part of CH2(CH2)6CH2, N(CH3)2), 2.34-2.52 (m, 8, part
of NCH2(CH2)6CH2, CH2CH3), 3.80 (t, J ) 6.6 Hz, 2, OCH2),
6.53 (d, J ) 8.8 Hz, 2, ArH ortho to OCH2), 6.72 (d, J ) 8.8
Hz, 2, ArH meta to OCH2), 6.98 (d, J ) 8.3 Hz, 2, ArH meta to
I), 7.09-7.21 (m, 5, PhH), 7.66 (d, J ) 8.5 Hz, 2, ArH ortho to
I); MS m/ z 581 (M+, 50). Anal. (C32H40NOI) C, H, N; I: calcd,
21.82; found 21.12.
(E)-1-[4-[[10-(N-Dim et h yla m in o)d ecyl]oxy]p h en yl]-1-
(4-iodoph en yl)-2-ph en yl-1-bu ten e (2i): white crystals (0.412
g, 68%); mp 70-72 °C; 1H NMR (CDCl3) δ 0.91 (t, J ) 7.4 Hz,
3, CH2CH3), 1.23-1.74 (m, 16, CH2(CH2)8CH2), 2.19-2.28 (m,
8, NCH2(CH2)5CH2, N(CH3)2), 2.67 (q, J ) 7.6 Hz, 2, CH2CH3),
3.80 (t, J ) 7.6 Hz, 2, OCH2), 6.53 (d, J ) 8.5 Hz, 2, ArH ortho
to OCH2), 6.72 (d, J ) 8.8 Hz, 2, ArH meta to OCH2), 6.98 (d,
J ) 8.4 Hz, 2, ArH meta to I), 7.11-7.18 (m, 5, PhH), 7.66 (d,
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