11β-substituted 13β-ethyl gonane derivatives exhibit reversal of antiprogestational activity

Article abstract of DOI:10.1016/S0039-128X(97)00119-0

The syntheses of three 17α-acetoxy-13β-ethyl-11β-aryl-18,19- dinorpregna-4,9-diene-3,20 diones from levonorgestrel are described. Despite their close structural similarity to the antiprogesterone CDB-2914, one of the compounds exhibits agonistic progestational activity, and the other two compounds are totally inactive.

Full text of DOI:10.1016/S0039-128X(97)00119-0

11␤-Substituted 13␤-ethyl gonane
derivatives exhibit reversal of
antiprogestational activity
Pemmaraju N. Rao,* James W. Cessac,* Richard P. Blye,† and Hyun K. Kim†
* Department of Organic Chemistry, Southwest Foundation for Biomedical Research,
San Antonio, Texas, USA; and † National Institute of Child Health and Human Development,
National Institutes of Health, Bethesda, Maryland, USA
The syntheses of three 17␣-acetoxy-13␤-ethyl-11␤-aryl-18,19-dinorpregna-4,9-diene-3,20 diones from levonorg-
estrel are described. Despite their close structural similarity to the antiprogesterone CDB-2914, one of the
compounds exhibits agonistic progestational activity, and the other two compounds are totally inactive.
(Steroids 63:50–57, 1998) © 1998 by Elsevier Science Inc.
Keywords: synthesis; 13␤-ethyl gonane derivatives; CDB-2914; progestin; antiprogestin
Introduction
Experimental
The discovery of mifepristone (Figure 1) as an antiproges-
tational agent prompted spectacular progress in the basic
understanding of progesterone and antiprogesterone action,
as well as in the development of improved analogs1 and
compounds for other therapeutic uses.2 Substitution at the
17-position by an ␣-acetoxy-␤-acetyl group yielded a com-
pound (CDB-2914, Figure 1) which exhibited antiprogesta-
tional and antiglucocorticoid activity similar to mifepris-
tone.3 Analogs with pure antiprogestational activity devoid
of antiglucocorticoid activity would be of great interest for
long-term use as contraceptive or therapeutic agents.
The increase in relative binding affinity (RBA) of
levonorgestrel (Figure 1) for the progestin receptor (PR)
over norethindrone4 (Figure 1), along with a concomitant
increase in progestational activity in rabbits,5 led us to
believe that a similar substitution of a 13␤-ethyl group for
the 18-methyl group of CDB-2914 could result in synthetic
analogs with increased antiprogestational activity. A prece-
dent for this type of modification in antiprogestins has been
reported in the literature with regard to the synthesis and
antiprogestational activity of a 13␤-ethyl analog (RU43855,
Figure 1)6 of mifepristone. In order to extend this investi-
gation into this type of structural modification, 13␤-ethyl
analogs of CDB-2914 with varying 11␤-aryl substituents
were synthesized and tested for biological activity.
Chemistry
Melting points were determined on a Thomas–Hoover apparatus
and are uncorrected. H NMR were determined in deuterochloro-
1
form either at 300 MHz using a General Electric QE-300 spec-
trometer or at 90 MHz using a Varian EM-390 spectrometer.
Infrared spectra were recorded on a Perkin–Elmer model 1600
FTIR instrument equipped with a diffuse reflectance accessory
using a KBr matrix. Optical rotations were measured using a
Rudolph Research Autopol II automatic polarimeter using a 1.0
dm cell. HPLC analysis was carried out on Waters Associates, Inc.
equipment monitored by a Lambda max Model 481 LC Spectro-
photometer. Mass spectral analyses(EI) were conducted by Dr.
Susan Weintraub of the University of Texas Health Science Center
at San Antonio using a Finnigan-MAT model 4615 mass spec-
trometer. Combustion analyses were performed by Midwest
MicroLabs Ltd., Indianapolis, Indiana. “Flash column” chroma-
tography was performed on 32–64 ␮M silica gel obtained from
Scientific Adsorbents Inc., Atlanta, Georgia. TLC Analyses were
carried out on silica gel GF (Analtech) glass plates (2.5 ϫ 10 cm
with 250 ␮M layer and prescored).
Most chemicals and solvents were analytical grade and used
without further purification. 4-Bromoacetophenone ethylene ketal
was prepared according to the procedure of Detty et al.7 Levonorg-
estrel 1 was obtained from Wyeth–Ayerst Research, Princeton,
New Jersey.
Ketalization of Levonorgestrel
A mixture of Levonorgestrel 1 (60 g, 192 mmol), ethylene glycol
(60 mL, 1.07 mol), p-toluenesulfonic acid monohydrate (3.0 g,
15.8 mmol) in benzene (1.75 L), was heated to reflux under
nitrogen in a flask equipped with a Dean–Stark trap for 16 h. The
solvent was removed in vacuo and the residue taken up in meth-
ylene chloride. The organic fractions were washed with saturated
Address reprint requests to Pemmaraju N. Rao, Department of Organic
Chemistry, Southwest Foundation for Biomedical Research, P.O. Box
760549, San Antonio, Texas 78245-0549.
Received June 27, 1997; accepted September 11, 1997.
Steroids 63:50–57, 1998
© 1998 by Elsevier Science Inc. All rights reserved.
655 Avenue of the Americas, New York, NY 10010
0039-128X/98/$19.00
PII S0039-128X(97)00119-0

11␤-Substituted 13␤-ethyl gonane derivatives: Rao et al.
Figure 1.
sodium bicarbonate solution (1ϫ), water (1ϫ) and brine (1ϫ),
combined, dried (Na2SO4), filtered, and concentrated in vacuo to
give the crude mixture of ⌬5- and ⌬5,(10)-isomers of the 3-ketal 2
(79 g). Analysis by HPLC (Waters Associates NovaPak C18;
CH3CN/H2O 1:1; 1 mL/min; ␭ ϭ 210 nm) indicated a 65:35 ratio
of the ⌬5,(10)-isomer to the ⌬5-isomer. 1H NMR (90 MHz) ␦ 0.93
(m, 13␤-CH2CH3), 2.57 (s, C'CH), 3.97 (m, ethylenedioxy
CH2’s), 5.47 (m, 6 C–H)ppm.
ϭ 20.7 Hz, C4-H2) ppm. IR (cmϪ1) 3402, 3301, 2939, 1727,
1698. MS (m/z): Mϩ ϭ 312. Analysis calculated for C12H28O2 ⅐
1/3 CH3CN: C, 79.79; H, 8.96. Found: C, 79.82; H, 8.86.
13␤-Ethyl-17␤-hydroxy-18,19-dinor-17␣-pregna-
4,9-dien-20-yn-3-one (5)
Under nitrogen, a solution of the ⌬5,(10) ketone 4 (36 g, 115.2
mmol) in dry pyridine (175 mL) was added to a solution of
pyridinium bromide perbromide (45 g, 126.6 mmol) in dry
pyridine (350 mL) preheated to 80°C. The reaction was stirred
at 85–90°C for 1 h, cooled to room temperature and then poured
into ice cold 2.5 N HCl solution (3 L). The crude product was
collected by filtration, washed well with water, and taken up in
ethyl acetate. The organic fractions were washed once with 1 N
HCl, once with water, and once with brine, then combined,
dried (Na2SO4), filtered and concentrated in vacuo. Crystalli-
zation of the residue from ether gave the pure diene 5 (21.1 g,
59%) in two crops. Concentration of the mother liquors fol-
lowed by Flash chromatography (3% acetone in CH2Cl2) and
crystallization from ether gave an additional 2.8 g of the pure
diene 5 (total yield ϭ 23.9 g, 66.8%). m.p. ϭ 124–126°C [lit.8
130°C]. 1H NMR (300 MHz) ␦ 1.067 (t, J ϭ 7.4 Hz, 13␤-
13␤-Ethyl-17␤-hydroxy-18,19-dinor-17␣-pregn-
5(10)-en-20-yn-3-one (4)
The ketal mixture 2 (79 g, assume 192 mmol) was dissolved in
glacial acetic acid (840 mL) and the system was flushed with
nitrogen. Water (360 mL) was added and the mixture was
heated to 50°C for 1 h. Heating was discontinued and the
mixture was stirred an additional 16 h. The reaction mixture
was diluted to 6 L with water and the precipitate was collected
by filtration and washed well with water. The solid was taken
up in CH2Cl2, filtered through Na2SO4 and concentrated in
vacuo to give 60.1 g solid residue. This material (3) was
separated on a large Flash chromatography column (14 ϫ 32
cm. bed of silica, 1.5% acetone in CH2Cl2 followed by 5%
acetone in CH2Cl2) into levonorgestrel 1 (24 g, 40% recovery)
and the ⌬5,(10)-isomer 4 (36 g, 60% from 1) both as white
crystalline solids. A small amount of the ⌬5,(10)-isomer 4 was
crystallized from acetonitrile/water for characterization. m.p. ϭ
169–171°C. 1H NMR (300 MHz) ␦ 0.975 (t, J ϭ 7.5 Hz,
13␤-CH2CH3), 2.607 (s, C'CH), 2.681 and 2.787 (each d, JAB
CH2CH3), 2.605 (s, C'CH), 5.685 (s, C4-H) ppm. IR (cmϪ1
)
3296, 2942, 1654, 1604. MS (m/z): Mϩ ϭ 310. Analysis cal-
culated for C21H26O2: C, 81.25; H, 8.44. Found: C, 81.28;
H, 8.45.
Steroids, 1998, vol. 63, January 51

Papers
Figure 2. Synthesis of 17␣-Acetoxy-
13␤-ethyl-11␤-aryl-18,19-dinorpregna-
3,20-diones.
mixture stirred until warming to room temperature. Solid
Hg(OAc)2 (0.8 g, 2.5 mmol) was added and the mixture stirred at
room temperature for ϳ10 min. This mixture was then added at
once to a flask containing the crude nitrate product 6 (11.7 g,
assume 32.2 mmol) and the reaction was stirred at room temper-
ature and monitored by TLC (5% acetone/CH2Cl2) which indi-
cated a complete reaction after 5.5 h. The reaction was then poured
into ice water (ϳ2 L) with stirring and the resulting precipitate was
collected by filtration, washed with water and taken up in ethyl
acetate. The organic fractions were washed with water (2ϫ), brine
(1ϫ), combined, dried (Na2SO4), filtered and concentrated in
vacuo. Purification of the residue by Flash chromatography (7%
acetone/CH2Cl2) followed by crystallization from ether gave the
formate ester 7 (3.3 g, 29% from 5) as a light brown solid. m.p. ϭ
156–158°C. 1H NMR (300 MHz) ␦ 0.731 (t, J ϭ 7.5 Hz, 13␤
CH2CH3), 2.197 (s, C21-H3), 5.706 (s, C4-H), 8.034 (s, formate)
ppm. IR (cmϪ1) 2936, 1726, 1664, 1613. MS (m/z): Mϩ ϭ 356.
13␤-Ethyl-17␤-hydroxy-18,19-dinor-17␣-pregna-
4,9-dien-20-yn-3-one 17-nitrate ester (6)
Under nitrogen, purified fuming nitric acid (5 mL, 113 mmol) was
added dropwise to freshly distilled acetic anhydride (11 mL, 116.6
mmol) cooled to 0°C in an ice bath. The mixture was then stirred
at 0°C for 10 min, room temperature for 30 min, then cooled again
to 0°C. A portion (14 mL, ϳ99 mmol AcONO2) of this solution
was added to a solution of the dienone 5 (10 g, 32.2 mmol) in dry
CH2Cl2 (30 mL) cooled to Ϫ15°C in an ice/solid NH4Cl bath. This
mixture was stirred at Ϫ15°C for 1 h then poured into a mixture of
ice/saturated sodium bicarbonate solution (ϳ400 mL). The mix-
ture was stirred until the ice had melted and then extracted with
CH2Cl2 (3ϫ). The organic fractions were washed with saturated
sodium bicarbonate solution (1ϫ), water (1ϫ) and brine (1ϫ),
then combined, filtered through Na2SO4 and concentrated in vacuo
to give 11.7 g residue as a yellow-brown foam. This material was
used directly in the subsequent reaction without further purifica-
tion. A small amount of material from a previous preparation was
purified by crystallization from ether for purposes of characteriza-
tion. m.p. ϭ 96–98°C. 1H NMR (300 MHz) ␦ 1.058 (t, J ϭ 7.4 Hz,
17␣-Hydroxy-13␤-ethyl-18,19-dinorpregna-4,9-
diene-3,20-dione (8)
13␤-CH2CH3), 2.741 (s, C'CH), 5.701 (s, C-4H) ppm. IR (cmϪ1
)
3294, 3234, 2946, 2112, 1659, 1633, 1296. MS (m/z): Mϩ ϭ 355.
Analysis calculated for C21H25NO4: C, 70.96; H, 7.09; N, 3.94.
Found: C, 70.73; H, 7.04; N, 3.88.
Under nitrogen, a 0.5 M solution of KHCO3 (200 mL, 100 mmol)
was added to a solution of the 17␣-formate 7 (25 g, 70.1 mmol) in
methanol (2.5 L). After stirring overnight at room temperature,
TLC (5% acetone/CH2Cl2) indicated ϳ80% reaction. An addi-
tional 50 mL of the 0.5 M KHCO3 solution was added and the
mixture heated to reflux for 1 h. At the end of that time, TLC (10%
acetone in CH2Cl2) indicated a complete reaction. The reaction
mixture was cooled to room temperature and glacial acetic acid
(3.2 mL, 55.7 mmol) was added. The methanol was removed in
17␣-Formyloxy-13␤-ethyl-18,19-dinorpregna-4,9-
diene-3,20-dione (7)
Under nitrogen, formic acid (99%, 80 mL) was added to dry DMF
(20 mL) cooled in an ice bath. The ice bath was removed and the
52 Steroids, 1998, vol. 63, January

11␤-Substituted 13␤-ethyl gonane derivatives: Rao et al.
vacuo and the suspended crude product in water was extracted
with CH2Cl2 (3ϫ). The organic fractions were washed with water
(2ϫ), dried (Na2SO4), filtered and concentrated in vacuo to give
23 g residue as an oil. Trituration of this material with ether gave
17 g 8 as a solid. Concentration of the mother liquors followed by
Flash chromatography (10% acetone in CH2Cl2) and ether tritura-
tion gave an additional 3 g of 8. Total amount of the 17␣-hydroxy
compound 8 obtained was 20 g (89%). m.p. ϭ 208–211°C. 1H
NMR (300 MHz) ␦ 0.717 (t, J ϭ 7.5 Hz, 13␤-CH2CH3), 2.357 (s,
C21-H3), 5.681 (s, C4-H) ppm. IR (cmϪ1) 3431, 2953, 1703, 1642,
1597. MS (m/z): Mϩ ϭ 328. Analysis calculated for C21H28O3: C,
76.79; H, 8.59. Found: C, 76.83; H, 8.59.
nitrogen. The system plus contents were flame dried under nitro-
gen. After cooling to room temperature, dry THF (25 mL) was
added followed by dibromoethane (ϳ0.1 mL) and the mixture was
stirred at room temperature. After evidence of reaction was ob-
served (bubbles), a solution of 4-bromo-N,N-dimethylaniline (5.72
g, 28.6 mmol) in dry THF (20 mL) was added via syringe. After
stirring at room temperature for 5–10 min, the reaction was initi-
ated and the reaction refluxed for about 20 min. The reaction
mixture was stirred for an additional 1 h during which time it had
cooled to room temperature. Solid copper (I) chloride (0.28 g, 2.8
mmol) was added with stirring followed 30 min later by a solution
of epoxide 10 (2.2 g, 5.08 mmol) in dry THF (10 mL). The
reaction was then stirred at room temperature for 1 h. Saturated
ammonium chloride solution (ϳ15 mL) was added dropwise to
quench the reaction. In order to oxidize Cu(I) to Cu(II), air was
drawn through the reaction mixture for 15 min via a 6 inch needle
inserted through the rubber septum and a slight vacuum applied to
the top of the reflux condenser. The reaction was then poured into
water (ϳ150 mL) and extracted with CH2Cl2 (3ϫ). The organic
fractions were washed with saturated NH4Cl solution (2ϫ), 2 N
NH4OH (2ϫ), water (1ϫ) and brine (1ϫ), then combined, filtered
through Na2SO4 and concentrated in vacuo to give 7 g blue-purple
residue as an oil. Purification via Flash chromatography (5%
acetone/CH2Cl2) followed by crystallization from ether gave 11a
3,20-bis-Ethylenedioxy-13␤-ethyl-17␣-hydroxy-
18,19-dinorpregna-4,9-diene (9)
Under nitrogen, a mixture of the 17␣-hydroxy compound 8 (18 g,
54.8 mmol), triethylorthoformate (23 mL, 138.1 mmol), ethylene
glycol (16 mL, 286 mmol), and toluenesulfonic acid monohydrate
(0.5 g, 2.6 mmol) in dry CH2Cl2 (300 mL) was stirred at room
temperature overnight. The mixture was diluted with CH2Cl2
(ϳ100 mL), washed with saturated sodium bicarbonate solution
(1ϫ), water (1ϫ), and brine (1ϫ). The organic fractions were
combined, dried (Na2SO4), filtered and concentrated in vacuo to
give 22 g residue as a brown solid. This material was crystallized
from CH2Cl2/MeOH containing a trace of pyridine to give diketal
9 (19.2 g, 84%). m.p. ϭ 110–113°C. 1H NMR (300 MHz) ␦ 0.893
(t, J ϭ 7.5 Hz, 13␤-CH2CH3), 1.392 (s, C21-H3), 3.986 (s over m,
ethylenedioxy CH2’s), 5.581 (m, C11-H)ppm. IR (cmϪ1) 3506,
2953, 2879. M/S (m/z): Mϩ ϭ 416. Analysis calculated for
C25H26O5 ⅐ 2/9 CH2Cl2: C, 69.57; H, 8.44. Found: C, 69.58;
H, 8.30.
1
(2.27 g, 78.8%). m.p. ϭ 137–140°C (d). H NMR (300 MHz) ␦
0.317 (t, J ϭ 7.5 Hz, 13␤-CH2CH3), 1.370 (s, C21-H3), 2.888 (s,
NMe2), 3.761–4.015 (m, ethylenedioxy CH2’s), 4.129–4.172 (m,
C11␣-H), 6.615 (d, J ϭ 39.6 Hz, J2 ϭ 8.7 Hz, aromatic C3Ј-H and
C5Ј-H), 7.044 (d, J ϭ 8.7 Hz, aromatic C2Ј-H and C6Ј-H) ppm. IR
(cmϪ1) 3530, 3471, 2949, 2884. MS (m/z): Mϩ ϭ 553. Analysis
calculated for C33H47NO6 ⅐ 1/2 H2O: C, 70.43; H, 8.60; N, 2.49.
Found: C, 70.40; H, 8.48; N, 2.54.
3,20-bis-Ethylenedioxy-13␤-ethyl-5␣,17␣-dihydroxy-
11␤-[4-(methylthio)phenyl]-18,19-dinorpregn-9-
ene (11b)
3,20-bis-Ethylenedioxy-13␤-ethyl-17␣-hydroxy-
5␣,10␣-epoxy-18,19-dinorpregn-9(11)-ene (10)
Hydrogen peroxide (30%, 5.6 mL, 54.66 mmol) was added to a
solution of hexafluoroacetone trihydrate (7.7 mL, 55.24 mmol) in
CH2Cl2 (150 mL) cooled to 0°C in an ice bath. The mixture was
stirred at 0°C for 30 min followed by the dropwise addition of
diketal 9 (18 g, 43.2 mmol) in CH2Cl2 (150 mL) cooled to 0°C
along with solid Na2HPO4 (8.6 g, 60.54 mmol). The mixture was
stirred at 0°C for 3 h, then overnight at 4°C. The reaction was
diluted with CH2Cl2 (ϳ150 mL) and washed with 10% Na2SO3
solution (2ϫ), and saturated sodium bicarbonate solution (1ϫ).
The organic fractions were combined, filtered through Na2SO4 and
concentrated in vacuo to give 19 g residue as a foam. Analysis by
TLC (2% acetone in CH2Cl2) indicated ϳ5–10% of starting ma-
terial remained. Analysis by NMR indicated epoxide mixture to
consist of a 65:35 mixture of the ␣- and ␤-isomers of the 5,10
epoxide. Trituration of this mixture with ether gave the pure solid
5␣,10␣-epoxide 10 (7.1 g, 38%). m.p. ϭ 145–147°C. 1H NMR
(300 MHz) ␦ 0.911 (t, J ϭ 7.5 Hz, 13␤-CH2CH3), 1.377 (s,
C21-H3), 3.80–4.10 (m, ethylenedioxy CH2’s), 6.043 (m, C11-H)
ppm. IR (cmϪ1) 3506, 2953, 2879. MS (m/z): Mϩ ϭ 432. Analysis
calculated for C21H36O6: C, 69.42; H, 8.39. Found: C, 69.29;
H, 8.49.
Following the same procedure used in the preparation of 11a, a
solution of 4-bromothioanisole (9.78 g, 48.15 mmol) in dry THF
(20 mL) was added to magnesium (1.17 g, 48.14 mmol), dibro-
moethane (ϳ0.1 mL), and a small crystal of I2 in dry THF (80
mL). After the reaction was initiated, the mixture was stirred at
ambient temperature for 1 h. Solid copper (I) chloride (0.475 g, 4.8
mmol) was added with stirring, followed 30 min later by a solution
of epoxide 10 (4.0 g, 9.6 mmol) in dry THF (20 mL). The reaction
was then stirred at room temperature for 2 h. Analysis by TLC
(10% acetone/CH2Cl2) of a small aliquot, quenched with saturated
ammonium chloride and extracted with ethyl acetate, indicated a
complete reaction at this time. Work up as previously described
gave 9.7 g crude product as a reddish oil. Trituration of this
material with pentane afforded compound 11b (4.135 g, 77%) as a
1
white solid. m.p. ϭ 182–183°C. H NMR (300 MHz) ␦ 0.280 (t,
J ϭ 7.5 Hz, 13␤-CH2CH3), 1.356 (s, C21-H3), 2.453 (s, SCH3),
3.737–4.015 (m, ethylenedioxy CH2’s), 4.201 (d, J ϭ 8.4 Hz,
C11␣-H), 7.122 (s, aromatic) ppm. IR (cmϪ1) 3535, 3502, 2941.
MS (m/z): Mϩ ϭ 556. Analysis calculated for C32H44O6S: C,
69.03; H, 7.97; S, 5.76. Found: C, 68.99; H, 7.94; S, 5.68.
3,20-bis-Ethylenedioxy-13␤-ethyl-5␣,17␣-dihydroxy-
11␤-[4-(N,N-dimethylamino)phenyl]-18,19-
dinorpregn-9-ene (11a)
3,20-bis-Ethylenedioxy-13␤-ethyl-5␣,17␣-dihydroxy-
11␤-{4-[1,1-(ethylenedioxy)ethyl]phenyl}-18,19-
dinorpregn-9-ene (11c)
Under anhydrous conditions, magnesium (0.76 g, 31.3 mmol) was
weighed into a 100 mL round bottom two-neck flask equipped
with a reflux condenser, a magnetic stir bar and a rubber septum.
A small crystal of I2 was added and the system flushed with dry
Following the same procedure used in the preparation of 11a, a
solution of 4-bromoacetophenone ethylene ketal1 (8.5 g, 34.97
mmol) in dry THF (20 mL) was added to magnesium (0.85 g,
34.97 mmol), 1,1-dibromoethane and a small crystal of I2 in dry
Steroids, 1998, vol. 63, January 53

Papers
THF (40 mL). After the reaction had initiated, the mixture was
stirred at ambient temperature for 2 h. Surprisingly, this Grignard
reagent is only partially soluble in THF, giving a suspension of
white solid upon complete formation. Solid copper (I) chloride
(0.346 g, 3.49 mmol) was added followed 30 min later by a
solution of epoxide 10 (3.0 g, 6.94 mmol) in dry THF (20 mL).
The reaction was complete after 1 h at room temperature. Work up
as previously described gave 9.5 g crude product mixture. Tritu-
ration of this material with ether afforded 2.5 g of the solid 11c.
Concentration of the filtrate in vacuo followed by Flash chroma-
tography (Et2O/CH2Cl2 1:1) gave an additional 0.7 g product.
Total yield of 11c was 3.2 g (77%). m.p. ϭ 182–183°C. 1H NMR
(300 MHz) ␦ 0.151 (t, J ϭ 7.5 Hz, 13␤-CH2CH3), 1.353 (s,
C21-H3), 1.634 (s, CH3 of 11␤-substituent), 3.698–4.038 (m,
ethylenedioxy CH2’s), 4.247 (d, J ϭ 8.4 Hz, C11␣-H), 7.175 (d,
J ϭ 8.4 Hz, aromatic C2Ј-H and C6Ј-H), 7.298 (d, J ϭ 8.4 Hz,
aromatic C3Ј-H and C5Ј-H) ppm. IR (cmϪ1) 3501, 2945. MS
(m/z): Mϩ ϭ 596. Analysis calculated for C35H48O8 ⅐ 1/6 H2O: C,
70.09; H, 8.12. Found: C, 70.19; H, 7.94.
13␤-Ethyl-11␤-(4-acetylphenyl)-17␣-hydroxy-18,19-
dinorpregna-4,9-diene-3,20-dione (12c)
Following the same procedure used to prepare compound 12a,
Grignard adduct 11c (3.2 g, 5.36 mmol) and 8.5% sulfuric acid (10
mL) were refluxed in ethanol (100 mL) for 1 h. After that time,
TLC (10% acetone in CH2Cl2) indicated a complete reaction. The
mixture was cooled to room temperature, diluted with water (ϳ100
mL) and neutralized with concentrated NH4OH solution. The
reaction mixture was further diluted with water (ϳ100 mL) and
extracted with ether (3ϫ). The organic fractions were washed with
water (2ϫ) and brine (1ϫ), then combined, dried (Na2SO4), fil-
tered and concentrated in vacuo to give 2.5 g residue as a foam.
This material was purified by Flash chromatography (Et2O/
CH2Cl2 1:1) followed by trituration with ether to give the 17␣-ol
12c (2.06 g, 86%). m.p. ϭ 151–154°C. 1H NMR (300 MHz)
␦–0.151 (t, J ϭ 7.2 Hz, 13␤-CH2CH3), 2.282 (s, C21-H3), 2.562
(s, 4Ј-acetyl CH3), 4.449 (d, J ϭ 8.1 Hz, C11␣-H), 5.755 (s,
C4-H), 7.293 (d, J ϭ 7.8 Hz, aromatic C2Ј-H and C6Ј-H), 7.853
(d, J ϭ 7.8 Hz, aromatic C3Ј-H and C5Ј-H)ppm. IR (cmϪ1) 3335,
2950, 1707, 1680, 1653, 1602. MS (m/z): Mϩ ϭ 446. Analysis
calculated for C29H34O4 ⅐ 2/3 H2O: C, 75.95; H, 7.77. Found: C,
76.01; H, 7.78.
13␤-Ethyl-11␤-[4-(N,N-dimethylamino)phenyl]-
17␣-hydroxy-18,19-dinorpregna-4,9-diene-3,20-
dione (12a)
17␣-Acetoxy-13␤-ethyl-11␤-[4-(N,N-dimethylamino)
phenyl]-18,19-dinorpregna-4,9-diene-3,20-
dione (13a)
Nitrogen was bubbled through a mixture of ethanol (50 mL) and
8.5% sulfuric acid (5 mL) for 30 min to remove oxygen. The
Grignard adduct 11a (2.27 g, 4.1 mmol) was added as a solid and
the mixture was refluxed for 1 h. After that time, TLC (10%
acetone/CH2Cl2) indicated complete reaction. The mixture was
cooled to room temperature, neutralized with saturated sodium
carbonate solution, diluted with water (ϳ100 mL) and extracted
with CH2Cl2 (3ϫ). The organic fractions were washed with satu-
rated sodium bicarbonate solution (1ϫ), water (1ϫ) and brine
(1ϫ), then combined, filtered through sodium sulfate, and concen-
trated in vacuo to give 2 g residue as a foam. This material was
purified by Flash chromatography (10% acetone/CH2Cl2) to give
Trifluoroacetic anhydride (8 mL, 56.6 mmol), glacial acetic acid
(3.3 mL, 57 mmol) and dry CH2Cl2 (50 mL) were combined and
stirred under nitrogen at room temperature for 30 min. Solid
p-toluenesulfonic acid monohydrate (0.5 g, 2.6 mmol) was added
and the mixture was cooled to 0°C in an ice bath. A solution of the
17␣-alcohol 12a (1.25 g, 2.8 mmol) in dry CH2Cl2 (10 mL) cooled
to 0°C was added and the mixture stirred at 0°C for 1 h. At the end
of that time, TLC (10% acetone in CH2Cl2) indicated complete
reaction. The reaction was carefully quenched with saturated po-
tassium carbonate solution (40 mL, ϳ200 mmol), diluted with
water (100 mL) and extracted with CH2Cl2 (3ϫ). The organic
fractions were washed with water (2ϫ) and brine (1ϫ), then
combined, filtered through Na2SO4 and concentrated in vacuo to
give 1.5 g residue as a foam. Purification by Flash chromatography
(5% acetone in CH2Cl2) followed by crystallization from CH2Cl2
1
the 17␣-ol 12a (1.27 g, 69.2%) as a foam. H NMR (90 MHz)
␦–0.07 (t, J ϭ 6.6 Hz, 13␤-CH2CH3), 2.27 (s, C21-H3), 2.89 (s,
NMe2), 4.35 (br. d, J ϭ 7.2 Hz, C11␣-H), 5.72 (s, C4-H), 6.60 (d,
J ϭ 9 Hz, aromatic C2Ј-H and C6Ј-H), 7.00 (d, J ϭ 9 Hz, aromatic
C3Ј-H and C5Ј-H) ppm. IR (cmϪ1) 3469, 2944, 2883, 1706, 1658,
1649, 1612. MS (m/z): Mϩ ϭ 447.
gave compound 13a (0.8 g, 57.6%). m.p. ϭ 233–236°C. [␣]D26
ϭ
ϩ210.73° (c ϭ 1.03, CHCl3). 1H NMR (300 MHz) ␦–0.057 (t, J ϭ
7.2 Hz, 13␤-CH2CH3), 2.116 (s, C21-H3 or OAc), 2.128 (s,
C21-H3 or OAc), 2.894 (s, NMe2), 4.367 (d, J ϭ 8.7 Hz, C11␣-H),
5.736 (s, C4-H), 6.608 (d, J ϭ 8.7 Hz, aromatic C2Ј-H and C6Ј-H),
13␤-Ethyl-11␤-[4-(methylthio)phenyl]-17␣-hydroxy-
18,19-dinorpregna-4,9-diene-3,20-dione (12b)
6.998 (d, J ϭ 8.4 Hz, aromatic C3Ј-H and C4Ј-H)ppm. IR (cmϪ1
)
Following the same procedure used to prepare compound 12a,
Grignard adduct 11b (3.79 g, 6.8 mmol) and 8.5% sulfuric acid (10
mL) were refluxed in ethanol (100 mL) for 45 min. After that time,
TLC (5% acetone in CH2Cl2) indicated a complete reaction. The
mixture was cooled to room temperature, diluted with water (ϳ100
mL) and neutralized with concentrated NH4OH solution. The
reaction mixture was further diluted with water (ϳ100 mL) and
extracted with ether (3ϫ). The organic fractions were washed with
water (2ϫ) and brine (1ϫ), then combined, dried (Na2SO4), fil-
tered and concentrated in vacuo to give 3.2 g residue as a yellow
foam. This material was purified by Flash chromatography (5%
acetone in CH2Cl2) followed by trituration with heptane to give the
17␣-ol 12b (1.95 g, 63.6%). m.p. ϭ 115–120°C. 1H NMR (90
MHz) ␦–0.10 (t, J ϭ 7.5 Hz, 13␤-CH2CH3), 2.27 (s, C21-H3),
2.43 (s, SMe), 4.37 (br. d, J ϭ 7.5 Hz, C11␣-H), 5.73 (s, C4-H),
7.13 (s, aromatic) ppm. IR (cmϪ1) 3461, 2941, 1706, 1648, 1591.
MS (m/z): Mϩ ϭ 450. Analysis calculated for C28H34O3S ⅐ 1/4
H2O: C, 73.89; H, 7.64; S, 7.04. Found: C, 73.94; H, 7.70; S, 6.88.
2943, 2878, 1730, 1707, 1654, 1610. MS (m/z): Mϩ ϭ 489.
Analysis calculated for C31H39NO4 ⅐ 1/4 H2O: C, 75.35; H, 8.06;
N, 2.83. Found: C, 75.22; H, 8.00; N, 2.84. Analysis by HPLC
(H2O/CH3CN/Et3N 30:70:0.033; Waters Associates NovaPak C18,
1 mL/min, ␭ ϭ 260 nm) indicated the product to be Ͼ99% pure
with a retention time of 7.8 min.
17␣-Acetoxy-13␤-ethyl-11␤-[4-(methylthio)phenyl]-
18,19-dinorpregna-4,9-diene-3,20-dione (13b)
Following the same procedure used to prepare compound 13a, the
17␣-alcohol 12b (1.7 g, 3.77 mmol) in dry CH2Cl2 (15 mL) was
reacted with the mixed anhydride prepared from trifluoroacetic
anhydride (15 mL, 106.4 mmol), glacial acetic acid (6.1 mL, 106.7
mmol) and p-toluenesulfonic acid monohydrate (1.0 g, 5.25 mmol)
in dry CH2Cl2 (75 mL) at 0°C. After stirring at 0°C for one hour,
TLC (10% acetone in CH2Cl2) indicated a complete reaction. The
54 Steroids, 1998, vol. 63, January

11␤-Substituted 13␤-ethyl gonane derivatives: Rao et al.
reaction was quenched at 0°C with concentrated NH4OH solution
(ϳ22 mL, 326 mmol), diluted with water (100 mL) and extracted
with CH2Cl2 (3ϫ). The organic fractions were washed with water
(2ϫ) and brine (1ϫ), then combined, filtered through Na2SO4 and
concentrated in vacuo to give 2.05 g residue as a yellow solid.
Purification via Flash chromatography (4% acetone in CH2Cl2)
followed by crystallization from ethyl acetate gave the acetate 13b
(1.13 g, 61%). m.p. ϭ 270–275°C (d). [␣]2D6 ϭ ϩ213.9° (c ϭ 1.01,
CHCl3). 1H NMR (300 MHz) ␦–0.103 (t, J ϭ 7.2 Hz, 13␤-
CH2CH3), 2.118 (s, C21-H3 or OAc), 2.131 (s, C21-H3 or OAc),
2.444 (s, SMe), 4.406 (d, J ϭ 8.1 Hz, C11␣-H), 5.757 (s, C4-H),
7.087 and 7.142 (each d, J ϭ 8.7 Hz, aromatic) ppm. IR (cmϪ1
2948, 1728, 1712, 1659, 1595. M/S (m/z): Mϩ ϭ 492. Analysis
calculated for C30H36O4S ⅐ 1/8 H2O: C, 72.80; H, 7.38; S, 6.48.
Found: C, 72.84; H, 7.41; S, 6.47. Analysis by HPLC (H2O/
CH3CN 3:7, Phenomenex Prodigy 5 ODS-2; 1 mL/min; ␭ ϭ 302
nm) indicated the product to be Ͼ99% pure with a retention time
of 7.2 min.
uterine horns were excised, cleaned of fat and connective tissue,
blotted on moist filter paper to express luminal fluid and weighed
to the nearest 0.1 mg. Midsections of each uterine horn (central to
the ligatures for intraluminal administration) were removed, fixed
in Bouin’s fluid, processed for sectioning at 5 microns and stained
with hematoxylin and eosin. Slides were read in a blinded fashion
and the degree of glandular proliferation was graded. Three dose
levels of unknown and standard were usually tested. Means and
standard error of the means were calculated and plotted on semilog
paper. Curve fitting, potency ratios and statistics were undertaken
using PROPHET.11–14
)
Antiprogestational assay. Antiprogestational activity was mea-
sured as the percent inhibition of the response to a standard
stimulating dose (0.8 mg total dose) of progesterone when both
substances were administered concomitantly using the protocol
described for progestational activity. Where compounds were ad-
ministered intraluminally, progesterone was injected subcutane-
ously for three days following estrogen priming. Treatment of data
was similar to that described above except that they are reported in
terms of percent inhibition.
17␣-Acetoxy-13␤-ethyl-11␤-(4-acetylphenyl)-18,19-
dinorpregna-4,9-diene-3,20-dione (13c)
Relative binding affinities for the progesterone
and glucocorticoid receptors
Following the same procedure used to prepare compound 13a, the
17␣-alcohol 12c (1.6 g, 3.58 mmol) in dry CH2Cl2 (10 mL) was
reacted with the mixed anhydride prepared from trifluoroacetic
anhydride (10 mL, 70.94 mmol), glacial acetic acid (4.1 mL, 71.7
mmol) and toluenesulfonic acid monohydrate (0.68 g, 3.6 mmol)
in dry CH2Cl2 (50 mL) at 0°C. After stirring at 0°C for 45 min,
TLC (10% acetone in CH2Cl2) indicated a complete reaction. The
reaction was quenched at 0°C with concentrated NH4OH solution
(ϳ15 mL, 222 mmol), diluted with water (100 mL) and extracted
with CH2Cl2 (3ϫ). The organic fractions were washed with water
(2ϫ) and brine (1ϫ), then combined, filtered through Na2SO4 and
concentrated in vacuo to give 1.6 g residue as a yellow solid.
Crystallization of this material from CH2Cl2/Et2O gave the acetate
13c (1.2 g, 68.5%) as a white solid. m.p. ϭ 268–270°C (d). [␣]D26
Uteri and Thymus glands were obtained from estradiol-primed
immature female rabbits of the New Zealand White strain. Tissues
were excised and immediately placed in ice cold TEGDM buffer
(10 mM Tris, pH 7.4; 1.5 mM EDTA; 10% glycerol vol/vol; 1 mM
dithiothreitol [DTT]; and 20 mM sodium molybdate). The tissues
were dissected free of connective tissue and fat, weighed, and
minced finely. Minced tissues were homogenized in three volumes
TEGDM/gm with four 10 s bursts of a VirTis Cyclone set at half
maximum speed with a 30 s cooling period (in ice) between bursts.
Homogenates were centrifuged at 109,663 ϫg at 4°C for 1 h to
yield the soluble cytosol fraction. Aliquots of cytosol were snap
frozen and stored at Ϫ75°C. All binding assays were carried out at
2–4°C for 16–18 h. The following radioactive ligands were used:
[1,2-3H(N)]-progesterone (50.0 Ci/mmol) for the progesterone re-
ceptor (PR) and [6,7-3H(N)]-dexamethasone (39.2 Ci/mmol) for
the glucocorticoid receptor (GR). For the progesterone receptor
RBA assays 0.02 mL uterine cytosol or TEGDM buffer, 0.05 mL
of various concentrations of test compounds or progesterone, 0.13
mL TEGDM buffer and 0.05 mL [3H]-progesterone were added to
duplicate tubes. For the glucocorticoid receptor RBA assays, 0.1
mL thymus cytosol or TEGDM buffer, 0.05 mL of various con-
centrations of test compounds or dexamethasone, 0.05 mL
TEGDM buffer and 0.05 mL [3H]-dexamethasone were added to
duplicate tubes. The concentrations of the test compounds, pro-
gesterone and dexamethasone ranged from 0.5 to 500 nM. Total
binding was measured at radioactive ligand concentrations of 3.5
nM and nonspecific binding was measured in the presence of a
200-fold molar excess of unlabeled progesterone (PR) or dexa-
methasone (GR), respectively.
In all incubations bound and free ligand concentrations were
separated using dextran-coated charcoal (DCC). A 0.1 mL aliquot
of DCC (0.5% charcoal/0.05% Dextran T-70) was added to each
tube. The tubes were vortexed and incubated on ice for 10 min.
Five tenths mL TEG buffer (without DTT or molybdate) was then
added to all tubes to improve supernatant recovery following
centrifugation. The charcoal was pelleted by centrifugation at
2,100 ϫg for 15 min at 4°C. The supernatants containing the
[3H]-steroid receptor complexes were decanted into vials contain-
ing 4 mL Optifluor (Packard Instrument Co.), vortexed, equili-
brated in a liquid scintillation counter for 30 min and then counted
for 2 min. This provided the quantity of receptor bound [3H]-
steroid at each competitor concentration.
1
ϭ ϩ184.4° (c ϭ 1.03, CHCl3). H NMR (300 MHz) ␦–0.164 (t,
J ϭ 7.5 Hz, 13␤-CH2CH3), 2.122 (s, C21-H3 or OAc), 2.138 (s,
C21-H3 or OAc), 2.570 (s, 4Ј-acetyl), 4.500 (d, J ϭ 8.7 Hz,
C11␣-H), 5.779 (s, 1 H, C4-H), 7.293 (d, J ϭ 8.4 Hz, aromatic
C2Ј-H and C6Ј-H), 7.863 (d, J ϭ 8.4 Hz, aromatic C3Ј-H and
C5Ј-H) ppm. IR (cmϪ1) 2951, 1729, 1712, 1683, 1660, 1596. MS
(m/z): Mϩ ϭ 488. Analysis calculated for C31H36O5: C, 76.20; H,
7.43. Found: C, 76.14; H, 7.33. Analysis by HPLC (H2O/CH3CN
3:7, Phenomenex Prodigy 5 ODS-2; 1 mL/min; ␭ ϭ 302 nm)
indicated the product to be Ͼ99% pure with a retention time of
4.1 min.
Biological assays
Progestational assay.9,10,11 Immature New Zealand White rabbits
approximately 1 kg in body weight were maintained under stan-
dard conditions of housing and allowed access to food and water
ad lib. The animals were randomized to groups of six rabbits each
and received daily subcutaneous injections of 5.0 ␮g estradiol
benzoate in sesame oil for six consecutive days. On the seventh
day, animals received the standard or test material dissolved in
sesame oil by subcutaneous or intraluminal injection. Subcutane-
ous injections were administered daily for five consecutive days.
Intraluminal injections of test compounds dissolved in sesame oil
were made into a 3–4 cm segment of one uterine horn and vehicle
only in the other, following Ketamine–Xylazine, 1:0.2 by weight,
(Ketasit Pitman Moore; Rompun Mobay Corp.) anesthesia. The
segment was ligated to prevent migration of test material or
vehicle. Animals were sacrificed 24 h after the last subcutaneous
injection and 3 days after intraluminal administration and the
Steroids, 1998, vol. 63, January 55

Papers
Table 1 Relative Binding Affinities and Biological Potency in the Rabbit
Relative binding affinity
Progestational
activityc
Antiprogestational
activityd
Compound
PRa
GRb
1
263
114
117
98
8.5
127
175
27
100
—
Inactive
12–21
Inactive
—
CDB-2914
13a
100
Inactive
Inactive
Inactive
13b
13c
159
18
a Progesterone ϭ 100%; immature estrogen-primed rabbit uterus.
b Dexamethasone ϭ 100%; immature estrogen-primed rabbit thymus.
c Progestational activity ϭ Clauberg: Oral administration to estrogen-primed immature rabbits; (Ϫ)-norgestrel (1) ϭ 100 (assigned).
d Antiprogestational activity ϭ Anti-Clauberg: Oral administration to estrogen-primed immature rabbits; CDB-2914 ϭ 100 (assigned).
The EC50 (Effective Concentration) for each standard curve
and each of the compound curves was determined by entering
the counting data (receptor bound [3H]-progesterone or [3H]-
pyridinium bromide perbromide in pyridine afforded the
dienone 5 in 67% yield. Conversion of 5 to the 17␤-nitrate
ester 6 followed by the mercuric acetate catalyzed isomer-
ization and hydrolysis procedure of Hofmeister et al17 gave
the formate ester 7 in 29% yield from compound 5. All of
the examples described by Hofmeister et al17 in their patent
consisted of ⌬4-3-keto, ⌬1,4-3-keto, or ⌬4,6-3-keto steroids
and the overall yields ranged from 19–65%. In our exper-
iments with the ⌬4,9-3-keto 5, the desired nitro derivative 6
was accompanied with other by-products, resulting in a
lower yield. Mild basic hydrolysis of the formate ester 7
gave the 17␣-hydroxy compound 8 in 89% yield. For the
introduction of the 11␤-aryl substituents, the general strat-
egy developed by Teutsch et al18 was adopted. Ketalization
of diene-dione 8 proceeded with a double bond shift to form
the 5(10), 9(11)-diene 9 in 84% yield. The regioselective
5,10-epoxidation of diene 9 was carried out using hexaflu-
oroacetone hydroperoxide generated in situ from hexaflu-
oroacetone trihydrate and hydrogen peroxide following the
general procedure of Teutsch et al.19 This technique was
modified by the addition of solid Na2HPO4 as buffer and
increasing the number of equivalents of hexafluoroacetone
trihydrate to 1.5. Application of this method to diene 9 gave
an epoxide mixture from which the pure 5␣,10␣-epoxide 10
could be isolated in 38% yield. The copper (I) catalyzed
addition of the Grignard reagent prepared from the ap-
propriate aryl bromide (4-bromodimethylaniline, 4-bromo-
thioanisole, and 4-bromoacetophenone ketal7) gave the cor-
dexamethasone) into
a four parameter sigmoidal computer
program (RiaSmart௢ Immunoassay Data Reduction Program,
Packard Instrument Co., Meridan, CT). The RBA for each test
compound was calculated using the following equation:
EC50 Standard
RBA ϭ
ϫ 100
EC50 Test Compound
where EC50 Standard ϭ molar concentration of unlabeled proges-
terone or dexamethasone required to decrease bound [3H]-
progesterone (PR) or [3H]-dexamethasone (GR) to 50% of the
respective buffer control (100% bound ligand) and EC50 Test
Compound ϭ molar concentration of test compound required to
decrease bound [3H]-progesterone (PR) or [3H]-dexamethasone
(GR) to 50% of the respective buffer control (100% bound ligand).
Results and discussion
Chemistry
Ketalization of levonorgestrel 1 (Figure 2) gave a quantita-
tive yield of the 3-ketal mixture 2, indicated by NMR and
HPLC to consist of a 65:35 ratio of the ⌬5(10) and the ⌬5(6)
isomers. Mild acidic hydrolysis of mixture 2 followed by
chromatographic separation of mixture 3 gave the ⌬5(10)
-
ketone 4 in 60% yield along with a 40% recovery of
levonorgestrel 1. Following the procedure of Perelman et
al,16 bromination-dehydrobromination of compound 4 using
Figure 3. Oral progestational
activity (Method of McPhail).
Legend: E (compound 13b); ‚
(compound 13c); ᮀ (compound
1); F (compound 13a); ■ (com-
pound 1). Straight regression
lines were computer fitted.
56 Steroids, 1998, vol. 63, January

11␤-Substituted 13␤-ethyl gonane derivatives: Rao et al.
responding Grignard adducts 11a–c in 77–79% yields. Acid
catalyzed hydrolysis and dehydration of compounds 11a–c
gave the analogous 17␣-hydroxy-diene-diones 12a–c in
yields ranging from 66 to 86%. Acetylation of compounds
12a–c was carried out in 58–69% yields following the
procedure of Carruthers and Garshasb20 to give the final
17␣-acetoxy products 13a–c.
References
1.
Teutsch G (1984). In: Agarwal MK (ed). Adrenal Steroid Antago-
nism. Walter de Gruyter and Co., Berlin, pp. 43–75.
Nieman LK, Chrousos GP, Kellner C, Spitz IM, Nisula BC, Cutler
GB, Merriam GR, Bardin CW, Loriaux DL (1985). Successful
treatment of Cushing’s syndrome with the glucocorticoid antagonist
RU 486. J Clin Endocrinol Metab 61:536–540.
2.
3.
Cook CE, Lee Y-W, Wani MC, Fail PA, Petrow V (1994). Effect of
D-ring substituents on antiprogestational (antagonist) and progesta-
tional (agonist) activity of 11␤-aryl steroids. Human Reprod 9:32–
39.
Biology
4.
5.
Moguilewsky M, Deraedt R (1981). Interactions between glucocor-
ticoid and progestin receptors. J Steroid Biochem 15:329–335.
Klimstra PD, Colton FB (1969). The chemistry of steroidal contra-
ceptives. In: Lednicer D (ed), Contraception: The Chemical Control
of Fertility. Marcel Dekker Inc., New York, pp. 69–159.
Teutsch G, Philibert D (1994). History and perspectives of antipro-
gestins from the chemist’s point of view. Human Reprod 9:12–31.
Detty MR, Murray BJ, Smith DL, Zumbulyadis N (1983). Cycliza-
tion of 3-(Arylchalcogeno)propenoyl Chlorides. 1. 1,2-Oxatelluro-
1-ium Halides via “Ipso” Acylation. J Am Chem Soc 105:875–882.
Joly R, Warnanat J, Joly J. 3-Methyl-3-alkoxy-4-oxagona-
5(10),9(11)-dienes. Fr., 1526961 680531; CA 73:15102.
McGinty DA, Anderson LP, McCollough NB (1939). Effect of local
application of progesterone on the rabbit uterus. Endocrinology
24:829–832.
Results of the in vitro relative binding affinities for proges-
terone and glucocorticoid receptors and in vivo Clauberg
and anti-Clauberg assays are shown in Table 1. Their rela-
tive binding affinities for the progesterone receptor in-
creased with the exception of 13b, and their relative binding
affinities for the glucocorticoid receptor decreased with the
exception of 13a. Oral administration of CDB-2914 and
levonorgestrel (1) were used as reference standards for the
anti-Clauberg and the Clauberg assay respectively. Surpris-
ingly, none of the 13␤-ethyl analogs displayed antiproges-
tational activity in the rabbit. On the contrary, one 13␤-ethyl
analog (13b) was found to exhibit agonistic activity in the
Clauberg assay. The 13␤-ethyl-11␤-(4-methylthiophenyl)-
analog (13b) has 12 to 21% progestational activity relative
to progesterone as shown in Table 1 and Figure 3. The
analogs 13a and 13c were devoid of oral progestational
activity (Method of McPhail, Figure 3). This is in contrast
with the marked antiprogestational activity reported for
most 11␤-aryl-19-norpregna-4,9-diene series3,6,21 and with
the abortive activity of RU43855, which is the 13␤-ethyl
analog of mifepristone previously reported by Teutsch and
Philibert.6 The latter has approximately 30% of the abortive
activity of mifepristone.6 No explanation for these results
can be given at the present time. However, the evidence
presented here suggests that the methyl portion of the an-
gular 13␤-ethyl group appears to play a role in the reversal
of antiprogestational activity of 13b and the loss of activity
of 13a and 13c in the rabbit. From the results shown in
Table 1, it appears that agonistic and antagonistic activities
are independent of relative binding affinity. A similar ob-
servation has been previously reported by Teutsch.22 Earlier
strategies23 for the design of antiprogestins centered around
mifepristone analogs targeted towards a high and specific
binding to the progesterone receptor. This strategy has be-
come of limited value.
6.
7.
8.
9.
10.
Clauberg C (1930). Zur Physiologie und Pathologie der Sexualhor-
mone, im besonderen Hormones des Corpus luteum. Zentralbl
Gynakol 54:2757–2770.
11.
12.
13.
14.
15.
16.
McPhail MK (1934). The assays of progestin. J Physiol 83:145–
156.
Bliss CI (1952). The Statistics of Bioassay. Academic Press, New
York.
Hollister C (1988). Prophet—A national computing resource for life
science research. Nuc Acids Res 16:1873–1875.
Holford N (1990). Drugmodel–Prophet Public Procedures. BBN
System and Technologies, Cambridge, MA.
Perry HM (1992). Prophet statistical and programming consultant
under Contract No. NO1-RR-2103.
Perelman M, Farkas E, Fornefeld EJ, Kraay RJ, Rapala RT (1960).
A new class of active steroids: The 19-nor-⌬4,9-3-ketosteroids. J Am
Chem Soc 82:2402–2403.
17.
18.
Hofmeister H, Annen K, Laurent H, Wiechert R. Process for the
preparation of pregnane derivatives. US Patent 4102908.
Belanger A, Philibert D, Teutsch G (1981). Regio and stereospecific
synthesis of 11␤-substituted 19-nor-steroids: Influence of 11␤-
substitution on progesterone receptor affinity. Steroids 37:361–382.
Teutsch G, Klich M, Bouchoux F, Cerede E, Philibert D (1994).
Synthesis of a fluorescent steroid derivative with high affinities for
the glucocorticoid and progesterone receptors. Steroids 59:22–26.
Carruthers NI, Garshasb S (1992). Synthesis of corticoids from
9␣-hydroxyandrost-4-ene-3,17-dione. J Org Chem 57:961–965.
Cook CE, Wani MC, Lee Y-W, Fail PA, Petrow V (1992). Reversal
of activity profile in analogs of the antiprogestin RU 486: Effect of
a 16␣-substituent on progestational (agonist) activity. Life Sci 12:
155–162.
19.
20.
21.
Currently, computer-aided drug design involving molec-
ular similarity methods24 and CoMFA (Comparative Mo-
lecular Field Analysis) technique25,26 using RBAs to pro-
gesterone and glucocorticoid receptors have already been
applied and resulted in the synthesis of more potent anti-
progestational agents than CDB-2914 with a decrease in
antiglucocorticoid activity. The results of these studies will
be reported in due course.
22.
Teutsch G (1985). Analogs of RU486 for the mapping of progestin
receptor: Synthetic and structural aspects. In: Baulieu EE, Segal SJ
(eds), The Antiprogestational Steroid RU486 and Human Fertility.
Plenum Press, New York, pp. 27–47.
23.
24.
Teutsch G, Gaillard-Moguilewsky M, Lemoine G, Nique F, Phili-
bert D (1991). Design of ligands for the glucocorticoid and proges-
tin receptors. Biochem Soc Trans 19:901–908.
Klebe G, Abraham U, Mietzner T (1994). Molecular similarity
indices in a comparative analysis (CoMSIA) of drug molecules to
correlate and predict their biological activity. J Med Chem 37:4130–
4146.
Acknowledgment
25.
26.
Cramer III RD, Patterson DE, Bunce JD (1988). Comparative mo-
lecular field analysis (CoMFA). 1. Effect of shape on binding of
steroids to carrier proteins. J Am Chem Soc 110:5959–5967.
Loughney DA, Schwender CF (1992). A comparison of progestin
and androgen receptor binding using the CoMFA technique. J Com-
put-Aided Mol Des 6:569–581.
This work was supported by Contract No. N01-HD-1-3137
from the National Institute of Child Health and Human
Development, National Institutes of Health, Bethesda,
Maryland.
Steroids, 1998, vol. 63, January 57