Ellipticine-Estradiol Conjugates
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 17 3371
(3.4 mL, 24 mmol), HOBt (1.22 g, 9 mmol), and N-[(tert-
butyloxycarbonyl)amino]-1,6-hexanediamine hydrochloride (10)
(1.90 g, 7.5 mmol) were added to a solution of 9 (1.98 g, 6 mmol)
in dry DMF (60 mL). The reaction mixture was cooled to -15
°C, and EDCI‚HCl (1.61 g, 8.4 mmol) was added in portions
as a solid. The resulting mixture was stirred at -15 °C for a
period of 4 h and slowly allowed to warm to room temperature.
After a period of 48 h, the reaction mixture was evaporated
under reduced pressure to dryness (oil pump). The residue
was dissolved in CH2Cl2 (250 mL) and washed with a saturated
solution of sodium bicarbonate (2 × 100 mL), water (100 mL),
and brine (100 mL), dried (MgSO4), and filtered. The filtrate
was flash chromatographed on silica gel, eluting with a
gradient of 25-40% EtOAc in hexanes to afford fractions
showing a single spot on TLC. These fractions were pooled
and evaporated under reduced pressure to dryness to afford
11 (2.3 g, 73%) as a white solid: mp 105-107 °C; TLC Rf )
Exp er im en ta l Section
Gen er a l. Melting points were determined in capillary
tubes on
a Thomas-Hoover Unimelt apparatus and are
uncorrected. 1H NMR spectra were recorded on Bruker ARX-
300 and Varian VXR-500 spectrometers. IR spectra were
recorded on a Perkin-Elmer 1600 FT-IR spectrophotometer.
Low-resolution chemical ionization mass spectra (CIMS) were
determined on a Finnigan 4000 spectrometer using 2-meth-
ylpropane as the reagent gas. High-resolution CIMS, low-
resolution fast atom bombardment mass spectra (FABMS), and
peak match FABMS were obtained on a Kratos MS50 spec-
trometer. Analytical thin-layer chromatography was done on
Whatman silica 60 K6F glass-coated plates with fluorescent
indicator. Column chromatography was performed using
Merck silica gel (230-400 mesh). Solvents were dried over 4
Å molecular sieves before using. Microanalyses were per-
formed at the Purdue Microanalysis Laboratory, and all values
are within (0.4% of the calculated compositions.
1
0.44 EtOAc/hexanes (3:1); H NMR (CDCl3, 300 MHz) δ 7.10
(d, 1 H, J ) 9 Hz), 6.70 (br s, 1 H), 6.64 (dd, 1 H, J ) 9 Hz, 3
Hz), 6.58 (d, 1 H, J ) 3 Hz), 4.57 (br s, 1 H), 3.24-3.37 (m, 2
H), 3.13 (m, 2 H), 2.77 (m, 2 H), 2.55 (d, 1 H, J ) 15 Hz), 2.34
(d, 1 H, J ) 15 Hz), 1.81-1.86 (br t, 4 H), 1.10-1.70 (m, 17
H), 1.44 (s, 9 H), 0.90 (s, 3 H); low-resolution FABMS m/ z
529 (MH+); high-resolution FABMS calculated MH+ 529.3641,
found 529.3652. Anal. (C31H48N2O5‚0.5H2O) C, H, N.
Eth yl 3,17â-Dih ydr oxy-1,3,5-estr atr ien e-17r-acetate (8).
A solution of ethyl bromoacetate (7) (10.81 g, 64.8 mmol) in
anhydrous toluene (15 mL) was added dropwise over a period
of 30 min to a mixture of activated zinc (4.0 g, 61.1 mmol) and
estrone (6) (5 g, 18.5 mmol) in anhydrous THF (60 mL), and
the resulting mixture was stirred at room temperature for 1
h. The mixture was ultrasonicated for a period of 6 h and then
warmed to 50 °C for a period of 2 h. After cooling to room
temperature, the reaction mixture was treated with 1%
sulfuric acid (150 mL) and then ethyl ether (100 mL) and
stirred for 20 min. The organic layer was separated, and the
aqueous layer was extracted with ethyl ether (2 × 100 mL).
The combined organic layers were washed with 10% NaHCO3
solution (100 mL), and brine (100 mL), dried (Na2SO4), and
filtered. The filtrate was evaporated under reduced pressure
to dryness, and the residue was flash chromatographed on
silica gel, eluting with 2:1, hexanes/ethyl ether. The fractions
showing a single spot corresponding to Rf ) 0.32 were pooled
and evaporated under reduced pressure to dryness to afford 8
(3.68 g, 56%) as a thick oil which slowly solidified on storing
at 5 °C to afford a white colored solid: mp 66-68 °C; TLC
(1:1 ethyl ether/hexanes) Rf ) 0.32; 1H NMR (CDCl3, 300 MHz)
δ 7.14 (d, 1 H, J ) 9 Hz), 6.62 (dd, 1 H, J ) 9.0 Hz, 3.0 Hz),
6.56 (d, 1 H, J ) 3 Hz), 4.96 (br s, 1 H), 4.39 (br s, 1 H), 4.21
(q, 2 H, J ) 6 Hz), 2.80-2.85 (m, 2 H), 2.67 (d, 1 H, J ) 18
Hz), 2.52 (d, 1 H, J ) 18 Hz), 2.24-2.31 (m, 1 H), 2.05-2.13
(m, 1 H), 1.60-1.98 (m, 5 H), 1.26-1.58 (m, 6 H), 1.31 (t, 3 H,
J ) 6 Hz), 0.94 (s, 3 H); low-resolution CIMS m/ z (%) 359
(MH+, 15), 341 (100); high-resolution CIMS calculated MH+
359.2222, found 359.2207.
3,17â-Dih yd r oxy-1,3,5-estr a tr ien e-17r-a cetic Acid (9).
A 10% aqueous solution of NaOH (20 mL) in ethanol (80 mL)
was added to a solution of 8 (3.13 g, 8.74 mmol), and the
resulting solution was stirred for 48 h at room temperature.
Most of the ethanol was removed under reduced pressure, and
the reaction mixture was diluted with water (150 mL). The
aqueous layer was extracted with ether (2 × 50 mL), and this
organic layer was discarded. The aqueous layer was then
acidified to pH 1 with 1 N HCl and reextracted with ether (3
× 100 mL). The combined organic layers were washed with
brine (100 mL), dried (Na2SO4), and filtered. The filtrate was
subjected to flash column chromatography on silica gel, eluting
first with EtOAc (250 mL) and then with 1% MeOH in EtOAc
to afford fractions showing a single spot on silica gel at Rf )
0.25 (CHCl3/MeOH, 9:1). These fractions were pooled and
evaporated to dryness under reduced pressure and then
coevaporated 3 times from a hexanes/ether mixture. This
residue was dried over CaSO4 under vacuum to afford 9 (2.31
g, 80%) as a white solid: mp 142-144 °C (lit.17 mp 142 °C);
TLC Rf ) 0.25 CHCl3/MeOH (9:1); 1H NMR (CDCl3, 300 MHz)
δ 7.13 (d, 1 H, J ) 9 Hz), 6.63 (dd, 1 H, J ) 9 Hz, 3 Hz), 6.57
(d, 1 H, J ) 3 Hz), 2.80-2.83 (m, 2 H), 2.76 (d, 1 H, J ) 15
Hz), 2.59 (d, 1 H, J ) 15 Hz), 2.28-2.32 (m, 1 H), 2.04-2.10
(m, 1 H), 1.27-1.94 (m, 11 H), 0.95 (s, 3 H); low-resolution
CIMS m/ z (relative intensity) 331 (MH+, 15), 313 (100); high-
resolution CIMS calculated MH+ 331.1909, found 331.1899.
N-(6-Am in oh exyl)-3,17â-h yd r oxy-1,3,5-estr a tr ien e-17r-
a ceta m id e (12). A solution of 11 (2.26 g, 4.28 mmol) in CH2-
Cl2 (100 mL) was cooled in an ice bath to 0-5 °C. Trifluoro-
acetic acid (15 mL) was added, and the deep yellow solution
was stirred at 0-5 °C for 6 h. Water (200 mL) was added
carefully to the reaction mixture and the organic layer
removed. The aqueous layer was basified to pH 8.5 with a
saturated solution of NaHCO3 and extracted with 10% MeOH
in CH2Cl2 (3 × 80 mL). The combined organic layers were
washed with brine (50 mL), dried (K2CO3), and filtered. The
filtrate was evaporated under reduced pressure to dryness and
flash chromatographed on silica gel, eluting first with 10%
MeOH in CHCl3 (250 mL), followed by 15% MeOH in CHCl3,
to afford fractions showing a single spot on TLC. These
fractions were pooled and evaporated under reduced pressure
to dryness to afford a brown oil. On drying this oil under
vacuum, a shiny brown hygroscopic solid formed which was
collected to afford 12 (1.65 g, 90%): mp softens and shrinks
at 85 °C; TLC Rf ) 0.15 CHCl3/MeOH/NH4OH (40:10:0.5); 1H
NMR (DMSO-d6, 300 MHz) δ 8.11 (t, 1 H, J ) 6 Hz), 7.03 (d,
1 H, J ) 9 Hz), 6.49 (dd, 1 H, J ) 9 Hz, 2 Hz), 6.42 (d, 1 H, J
) 2 Hz), 3.05 (br apparent q, 2 H, J ) 6 Hz), 2.67-2.71 (m, 2
H), 2.56 (t, 2 H, J ) 6 Hz), 2.22-2.27 (m, 3 H), 2.03 (m, 1 H),
1.50-1.85 (m, 5 H), 1.10-1.45 (m, 15 H), 0.80 (s, 3 H); low-
resolution CIMS m/ z (relative intensity) 429 (MH+, 10), 411
(20), 271 (40), 159 (100); high-resolution CIMS calculated MH+
429.3117, found 429.3112. Anal. (C26H40N2O3‚0.5CF3COOH)
C, H, N.
2-[(p -Nit r op h en oxy)ca r b on yl]-1,2-d ih yd r oellip t icin e
(14). A suspension of ellipticine (1) (0.246 g, 1 mmol) in
anhydrous THF (11 mL) was cooled to -70 °C in a dry ice/
acetone bath, and a solution of p-nitrophenyl chloroformate
(13) (0.242 g, 1.2 mmol) in THF (5 mL) was added slowly to
this suspension. There was immediate formation of a red
precipitate. After stirring for 5 min, a 1.0 M solution of
NaCNBH3 in THF (2.5 mL, 2.5 mmol) was added slowly via a
syringe drive and the mixture stirred for 10 min at -70 °C.
The mixture was poured into ice/water (100 mL) and extracted
with CHCl3 (2 × 80 mL). The combined CHCl3 extracts were
washed with 1 N HCl (80 mL) and cold water (80 mL), dried
(MgSO4), and filtered. The filtrate was evaporated under
reduced pressure to afford 14 (0.41 g, 99%) as an orange-
colored solid: mp 233-234 °C (dec.); TLC Rf ) 0.62 EtOAc/
hexanes (1:1); 1H NMR (300 MHz, DMSO-d6) δ 11.19 (s, 1 H),
8.33 (d, 2 H, J ) 9 Hz), 8.14 (dd, 1 H, J ) 8, 9 Hz), 7.63 and
7.58 (2 d, 2 H, J ) 9 Hz), 7.51 (d, 1 H, J ) 9 Hz), 7.37 (dd, 1
H, J ) 7, 8 Hz), 7.15 (dd, 1 H, J ) 7, 8 Hz), 6.97 and 6.94 (2
d, 1 H, J ) 9 Hz), 6.36 (d, 1 H, J ) 9 Hz), 5.22 and 5.02 (2 s,
2 H), 2.71 (s, 3 H), 2.51 (s, 3 H); low-resolution CIMS m/ z
(relative intensity) 414 (MH+, 100), 247 (40), 140 (20); high-
resolution CIMS calculated MH+ 414.1454, found 414.1441.
N-[6-[(ter t-Bu tyloxyca r bon yl)a m in o]h exyl]-3,17â-d ih y-
dr oxy-1,3,5-estr atr ien e-17r-acetam ide (11). Triethylamine