Fan et al.
“Characterization of Molecular and Structural Determinants of Selective Estrogen
Receptor Downregulators”
Supplemental Information
Synthetic methods
All reagents were purchased from Sigma-Aldrich. Proton and
13
C nuclear magnetic
resonance spectra (1H NMR, 13C NMR) were obtained on a Bruker ARX300 (300 MHz)
instrument; 1H NMR chemical shifts are reported as δ values in parts per million (ppm)
downfield from internal tetramethylsilane.
13
C NMR chemical shifts are reported as δ
values with reference to the solvent peak. Mass spectrometry (MS) and NMR instruments
were provided by the Shared Resource center of the Purdue Cancer Center.
Synthesis of 7604 analogs
(E)-4-(1-(4-methoxyphenyl)-2-phenylbut-1-enyl)benzaldehyde.
Activated
magnesium
turnings (0.75 g, 31 mmol) were dissolved in tetrahydrofuran (30 mL) and 4bromobenzaldehyde diethyl acetal (6.25 mL, 31 mmol) was added dropwise. The reaction
was heated gently to initiate the reaction and was then stirred at room temperature for 6
hours until the magnesium was consumed. The reaction was them cooled to 0 ˚C and 1(4-methoxyphenyl)-2-phenylbutan-1-one (6.0 g, 23.8 mmol) [1] dissolved in THF (5
mL) was added. The reaction stirred at room temperature for 3 hours and saturated
aqueous NH4Cl solution (20 mL) and water (20 mL) were then added. The solution was
then extracted with diethyl ether (3 x 25 mL) and the organic layer was concentrated to a
yellow oil. The yellow oil was then dissolved in ethanol (50 mL) and concentrated
hydrochloric acid (10 mL) was added. The solution was heated to reflux for 2 hours,
cooled to room temperature and then the ethanol was removed under reduced pressure.
Water (20 mL) was then added to the remaining aqueous solution and the compound was
extracted with dichloromethane (3 x 30 mL). The organic layer was dried with MgSO 4
and concentrated to a brown oil. After purification by flash silica gel chromatography
using 4:1 hexane: ethyl acetate as the eluent. The title compound (6.2 g, 18.1 mmol) was
isolated as a single stereoisomer in 76% yield. Spectroscopic information matched
previous reports [2].
(E)-4-(4-((E)-1-(4-hydroxyphenyl)-2-phenylbut-1-enyl)phenyl)but-3-en-2-one (7604-ket)
A solution of potassium bis(trimethylsilyl)amide (0.5 M in toluene) (14 mL, 7.0 mmol)
was added to a stirring 0 ˚C solution of diethyl (2-oxopropyl)phosphonate (1.35 g, 7.0
mmol) in tetrahydrofuran (25 ml). After stirring for 15 min at 0 ˚C, the solution was
cooled to -78 ˚C, and a solution of (E)-4-(1-(4-methoxyphenyl)-2-phenylbut-1enyl)benzaldehyde (7) (2.0 g, 5.8 mmol) in tetrahydrofuran (25 ml) was added dropwise.
The reaction was then allowed to warm to room temperature and stirred 16 h overnight.
The solution was poured into saturated sodium chloride, extracted with ethyl acetate,
dried over anhydrous magnesium sulfate, and solvent removed under reduced pressure to
give a yellow oil. The yellow oil was then dissolved in dichloromethane (30 mL) and
cooled to 0 ˚C. Boron tribromide (1 M in CH2Cl2) (18 mL, 18 mmol) was then added
dropwise and the solution stirred for 2 hours at 0 ˚C. Methanol (10 mL) was then added
and allowed to stir for 30 minutes, followed by the addition of water (20 mL). The layers
were then separated and the aqueous layer was then also extracted three times with
dichloromethane (30 mL). The organic layers were then combined, dried with MgSO4
and concentrated. After purification by flash silica gel chromatography using 9:1
methylene chloride: methanol as the eluent, the title compound (1.2 g, 3.13 mmol) was
isolated in 54% yield. Spectroscopic information matched previous reports [3].
(E)-3-(4-((E)-1-(4-hydroxyphenyl)-2-phenylbut-1-enyl)phenyl)acrylic acid (GW7604) A
solution of potassium bis(trimethylsilyl)amide (0.5 M in toluene) (14 mL, 7.0 mmol) was
added to a stirring 0 ˚C solution of trimethyl phosphonoacetate (1.27 g, 7.0 mmol) in
tetrahydrofuran (25 ml). After stirring for 15 min at 0 ˚C, the solution was cooled to -78
˚C, and a solution of (E)-4-(1-(4-methoxyphenyl)-2-phenylbut-1-enyl)benzaldehyde (7)
(2.0 g, 5.8 mmol) in tetrahydrofuran (25 ml) was added dropwise. The reaction was then
allowed to warm to room temperature and stirred 16 h overnight. The solution was
poured into saturated sodium chloride, extracted with ethyl acetate, dried over anhydrous
magnesium sulfate, and solvent removed under reduced pressure to give a yellow oil. The
yellow oil was then dissolved in ethanol (20 mL) and THF (20 mL) and then aqueous 1
M KOH (20 mL) was added. The solution was heated to reflux for 2 hours, cooled to
room temperature and acidified with concentrated HCl (2 mL) and then extracted three
times with methylene chloride (20 mL). The organic layers were combined, dried with
MgSO4 and concentrated to a yellow solid.
The yellow solid was then dissolved in dichloromethane (30 mL) and cooled to 0
˚C. Boron tribromide (1 M in CH2Cl2) (18 mL, 18 mmol) was then added dropwise and
the solution stirred for 2 hours at 0 ˚C. Methanol (10 mL) was then added and allowed to
stir for 30 minutes, followed by the addition of water (20 mL). The layers were then
separated and the aqueous layer was then also extracted three times with dichloromethane
(30 mL). The organic layers were then combined, dried with MgSO4 and concentrated.
After purification by flash silica gel chromatography using 9:1 methylene chloride:
methanol as the eluent, the title compound (0.75 g, 1.97 mmol) was isolated in 37% yield.
Spectroscopic information matched previous reports [4].
(E)-3-(4-((E)-1-(4-hydroxyphenyl)-2-phenylbut-1-enyl)phenyl)acrylamide (7604-NH2)
(E)-3-(4-((E)-1-(4-hydroxyphenyl)-2-phenylbut-1-enyl)phenyl)acrylic acid (0.2 g, 0.54
mmol) was dissolved in DMF (3 mL) and N-hydroxybenzotriazole (87 mg, 0.65 mmol),
1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide (125 mg, 0.65 mmol) and triethylamine
(0.1 mL) were added. Concentrated aqueous ammonium hydroxide (0.5 mL) was added
and the solution was placed in a sealed tube and allowed to react for 16 hours. Saturated
aqueous ammonium chloride solution (10 mL) was then added and the solution was
extracted three times with methylene chloride (10 mL). The organic layers were
combined, dried with MgSO4 and concentrated. After purification by flash silica gel
chromatography using 9:1 methylene chloride: methanol as the eluent, the title compound
(0.16 g, 0.43 mmol) was isolated in 80% yield. 1H NMR (CD3OD) of 1:1 E:Z mixture δ
0.83 (t, J = 7.2 Hz, 3 H), 2.39 (q, J = 7.3 Hz, 2 H), 6.32 (d, J = 2.6 Hz, 1 H), 6.37 (d, J =
15.8 Hz, 1 H), 6.52 (s, 1 H), 6.56 (d, J = 8.9 Hz, 1 H), 6.68 (d, J = 8.6 Hz, 1 H), 6.78 (d, J
= 8.6 Hz, 2 H), 6.93 (d, J = 8.6 Hz, 1 H), 6.99-7.17 (m, 7 H), 7.28 (d, J = 15.8 Hz, 1 H),
7.45 (s, 1H) 7.46 (d, J = 2.6 Hz, 1 H)
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