Synthesis, characterization and biological evaluation of some 16E-arylidene androstane derivatives as potential anticancer agents

Article abstract of DOI:10.1016/j.steroids.2011.03.015

A series of new 16E-arylidene androstane derivatives were synthesized and characterized. The new compounds were screened for their anticancer activities against the human cancer cell lines SW480, A549, HepG2 and HeLa in vitro using the MTT assay. The resu

Full text of DOI:10.1016/j.steroids.2011.03.015

Steroids 76 (2011) 709–723
Contents lists available at ScienceDirect
Steroids
journal homepage: www.elsevier.com/locate/steroids
Synthesis, characterization and biological evaluation of some 16E-arylidene
androstane derivatives as potential anticancer agents
Hao Guoa, Haotian Wua, Jin Yanga, Yuling Xiaoa, Hans-Josef Altenbachb, Guofu Qiua,
Hao Hua, Zhongyuan Wua, Xianran Hea, Dingshan Zhoua, Xianming Hua,∗
a State Key Laboratory of Virology, College of Pharmacy, Wuhan University, Wuhan 430072, China
b Bergische Universität Wuppertal, FB 9 – Organische Chemie, Gauss-Str. 20, 42097 Wuppertal, Germany
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of new 16E-arylidene androstane derivatives were synthesized and characterized. The new com-
pounds were screened for their anticancer activities against the human cancer cell lines SW480, A549,
HepG2 and HeLa in vitro using the MTT assay. The results of the in vitro study showed that a number of
compounds have shown IC50 values lower than 20 ␮M against the four cancer cell lines.
© 2011 Elsevier Inc. All rights reserved.
Received 14 February 2011
Received in revised form 19 March 2011
Accepted 26 March 2011
Available online 5 April 2011
Keywords:
16E-arylidene androstane
Heterocycles
Synthesis
Anticancer activity
1. Introduction
It has been reported that some tertiary amino steroids, 2␤,16␤-
dipiperidino-5␣-androstane-3␣,17␤-diol dipivalate (DAP) (Fig. 1)
Cancer is presently the second most important disease leading
to death in both developing and developed countries according
to the WHO. Among common currently available drugs for the
treatment of cancer are bioalkylating agents (chlormethine) [1,2],
antimetabolic agents (fluorouracil) [3,4], anticancer antibiotics
(doxorubicin) [5], and compounds from plants and their derivatives
(paclitaxel) [6]; however, a dose of anticancer drug sufficient to kill
tumor cells is often toxic to normal tissue, thus leading to numerous
side effects which, in turn, limit treatment efficacy. Long term effec-
tiveness is often limited by dose-related cumulative cardiotoxicity
and development of acquired drug resistance [7–9]. Therefore, the
new therapeutic agents for the treatment of cancer thus needed
should be more active and produce less side effects.
Steroids elicit diverse biological action via various functional
groups located around the periphery of their rigid tetracyclic core.
They have the ability to regulate a variety of biological processes
and thus are potential drug candidates for the treatment of a large
number of diseases including breast cancer [10], prostate cancer
[11], leukaemia [12,13], autoimmune diseases [14], osteoporosis
[15], etc. Most steroid based pharmaceuticals are semi-synthetic
compounds, or analogues, developed from the lead compound and
prepared by connecting special functional groups to the core struc-
ture of the steroid.
[16] exhibit significant antitumor activity against transplantable
tumors in three animal species including mouse, hamster, and rat.
The acute toxicity of DAP was similar to that of cyclophosphamide
and considerably lower than vinblastine. In an effort to find the
new analogues of this type of novel steroid molecule that exhibit
higher activity along with simpler chemical reactions and moreover
are easily prepared, some 16E-arylidene androstene derivatives
have emerged as potent anticancer agents, such as compound DPJ-
RG-1110 [17,18], compound I [19] and compound II [20] (Fig. 1).
These previous studies indicate that incorporation of a hetero-
cyclic ring in the steroid backbone demonstrated an important
impact on the activity. In addition, a substitution of the phenyl
ring at the C-16 position also effects the activity. This type of 16E-
arylidene androstene-aminosteroid derivative represents a novel
class of potential anticancer agents.
In order to elucidate the structure–activity relationship for this
type of compound we studied the synthesis and cytotoxic activity
of 16E-arylidene androstane-3-aminosteroid derivatives with dif-
ferent substituents and we obtained additional information on the
potential pharmacophoric core.
2. Experimental
2.1. General
∗
All the chemicals and solvent were purchased from commercial
sources. Solvents and reagents were dried and purified according
Corresponding author. Tel.: +86 27 68759887; fax: +86 27 68759850.
E-mail address: cassie gh@hotmail.com (X. Hu).
0039-128X/$ – see front matter © 2011 Elsevier Inc. All rights reserved.
doi:10.1016/j.steroids.2011.03.015

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H. Guo et al. / Steroids 76 (2011) 709–723
2.2.4. 16-(3,4,5-trimethoxy-benzylidene)-5˛-androstan-3ˇ-ol-
17-one (2d)
White solid. Yield 91%. m.p. 130–132 ◦C; 1H NMR (400 MHz,
CDCl3): 0.87 (s, 3H, CH3-19), 0.97 (s, 3H, CH3-18), 3.57–3.63 (m,
1H, C3-OH), 3.89 (s, 9H), 7.35 (s, 1H, CH), 6.77 (s, 2H). 13C NMR
(150 MHz, CDCl3): ı 12.32, 14.18, 20.54, 22.68, 28.37, 29.35, 31.12,
31.41, 31.64, 34.67, 35.73, 36.84, 38.02, 44.82, 47.60, 49.51, 54.48,
56.15, 60.96, 71.10, 107.62, 131.15, 133.10, 135.27, 139.23, 153.16,
209.75. Anal. Calcd. for C29H40O5: C 74.33, H 8.60; Found: C 74.51,
H 8.42.
2.2.5. 16-(4-chloro-benzylidene)-5˛-androstan-3ˇ-ol-17-one
(2e)
Fig. 1. Structures of DAP and DPJ-RG-1110.
White solid. Yield 91%. m.p.171–173 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.89 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 3.59–3.63 (m,
1H, C3-OH), 7.39 (s, 1H, CH), 7.38 (d, 2H, J = 8.56 Hz), 7.47 (d, 2H,
J = 8.48 Hz). 13C NMR (150 MHz, CDCl3): ı 12.35, 14.49, 20.56, 21.06,
28.39, 29.27, 31.13, 31.45, 31.65, 34, 73, 35.75, 36.87, 38.05, 44.84,
47.64, 49.46, 54.52, 60.41, 71.15, 128.94, 131.42, 134.13, 135.08,
136.67, 209.64. Anal. Calcd. for C26H33ClO2: C 75.61, H 8.05; Found:
C 75.62, H 8.00.
to the literature methods. Melting points were determined on a
XT-4 apparatus (uncorrected). 1H NMR and 13C NMR spectra were
obtained on a Varian Mercury VX400 apparatus in CDCl3 with TMS
as internal standard. Mass spectra were recorded on a Shimadzu
LCMS-2020 spectrometer. X-ray diffraction data were collected on
Bruker SMART APEX CCD diffractometer. Elemental analysis was
carried out on a VarioEL III (German) instrument. Silica gel was
used for analytical and flash chromatography.
2.2.6. 16-(4-bromo-benzylidene)-5˛-androstan-3ˇ-ol-17-one
(2f)
2.2. General procedure for the synthesis of compounds 2a–h
(Scheme 1)
White solid. Yield 91%. m.p. 182–184 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.89 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 3.59–3.65 (m,
1H, C3-OH), 7.36 (s, 1H, CH), 7.39 (d, 2H, J = 8.52 Hz), 7.54 (d, 2H,
J = 8.44 Hz). 13C NMR (150 MHz, CDCl3): ı 12.35, 14.21, 14.49, 20.56,
21.09, 28.38, 29.29, 31.12, 31.45, 31.63, 34.72, 35.74, 36.86, 38.04,
44.83, 47.66, 49.44, 54.50, 60.43, 71.16, 123.44, 131.65, 134.54,
136.84, 209.66. Anal. Calcd. for C26H33BrO2: C 68.27, H 7.27; Found:
C 68.51, H 7.08.
To a stirred solution of epiandrosterone (1.45 g, 5.00 mmol)
and sodium hydroxide (1.00 g, 25.00 mmol. 5.0 equiv) in methanol
was added the corresponding aldehyde (7.50 mmol, 1.5 equiv). The
resulting solution was stirred for 12 h at 40 ◦C until the reaction
was completed (monitored by TLC). Cold water was added to the
reaction mixture and the precipitate obtained was filtered, washed
with water, dried and crystallized from methanol to yield 2a–h.
2.2.7. 16-(4-fluoro-benzylidene)-5˛-androstan-3ˇ-ol-17-one
(2g)
2.2.1. 16-(4-methyl-benzylidene)-5˛-androstan-3ˇ-ol-17-one
(2a)
White solid. Yield 95%. m.p. 181–183 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.89 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 3.59–3.66 (m,
1H, C3-OH), 7.40 (s, 1H, CH), 7.12 (t, 2H), 7.54 (t, 3H). 3C NMR
(150 MHz, CDCl3): ı 12.35, 14.52, 20.56, 28.40, 29.20, 31.14, 31.45,
31.64, 34.72, 35.74, 36.86, 38.04, 44.84, 47.62, 49.50, 54.52, 71.15,
115.93, 131.73, 132.21, 135.69, 161.76, 164.25, 209.80. Anal. Calcd.
for C26H33FO2: C 78.75, H 8.39; Found: C 78.61, H 8.22.
White solid. yield 91%. m.p. 165–167 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.89 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 3.59–3.65 (m,
1H, C3-OH), 7.42 (s, 1H, CH), 7.22 (d, 2H, J = 7.92 Hz), 7.44 (d,
2H, J = 8.04 Hz), 2.40 (s, 3H, Ph-CH3). 13C NMR (150 MHz, CDCl3):
ı 12.35, 14.55, 20.59, 21.48, 28.43, 29.35, 31.13, 31.44, 31.68, 34.72,
35.74, 36.87, 38.05, 44.85, 47.59, 49.57, 54.53, 60.43, 71.13, 129.44,
130.36, 132.85, 135.19, 139.55, 210.09. Anal. Calcd. for C27H36O2:
C 82.61, H 9.24; Found: C 82.34, H 9.31.
2.2.8. 16-(furan-2-ylmethylene)-5˛-androstan-3ˇ-ol-17-one
(2h)
2.2.2. 16-(4-methoxy-benzylidene)-5˛-androstan-3ˇ-ol-17-one
(2b)
White solid. Yield 95%. m.p. 154–156 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.88 (s, 3H, CH3-19), 0.93 (s, 3H, CH3-18), 3.60–3.65 (m,
1H, C3-OH), 7.21 (s, 1H, CH), 6.52 (q, 1H), 6.66 (d, 1H, J = 3.44 Hz),
7.57 (d, 1H, J = 1.68 Hz). 13C NMR (150 MHz, CDCl3): ı 12.35, 14.62,
20.59, 28.44, 28.91, 31.14, 31.44, 31.62, 34.69, 35.74, 36.87, 38.05,
44.87, 47.77, 49.11, 54.55, 71.15, 112.32, 115.51, 119.51, 133.68,
144.66, 152.27, 209.94. Anal. Calcd. for C24H32O3: C 78.22, H 8.75;
Found: C 78.19, H 8.81.
White solid. yield 94%. m.p. 156–158 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.89 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 3.59–3.61 (m, 1H
C3-OH), 3.84 (s, 3H, OCH3), 7.38 (s, 1H, CH), 6.92 (d, 2H, J = 8.76 Hz),
7.49 (d, 2H, J = 8.76 Hz). 13C NMR (150 MHz, CDCl3): 12.35, 14.58,
20.59, 28.44, 29.28, 31.15, 31.44, 31.69, 34.72, 35.74, 36.87, 38.05,
44.86, 47.53, 49.63, 54.55, 55.36, 71.13, 114.19, 128.36, 132.07,
132.76, 133.77, 160.41, 210.13. Anal. Calcd. for C27H36O3: C 79.37,
H 8.88; Found: C 79.43, H 9.03.
2.3. General procedure for the synthesis of compounds 3a–h
2.2.3. 16-(4-nitro-benzylidene)-5˛-androstan-3ˇ-ol-17-one (2c)
Yellow solid. Yield 93%. m.p. 258–260 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.90 (s, 3H, CH3-19), 0.99 (s, 3H, CH3-18), 3.60–3.63 (m,
1H, C3-OH), 7.45 (s, 1H, CH), 7.67 (d, 2H, J = 8.80 Hz), 8.27 (d, 2H,
J = 8.84 Hz). 13C NMR (150 MHz, CDCl3): 12.33, 14.43, 20.51, 28.33,
29.38, 31.10, 31.39, 31.60, 34.72, 35.74, 36.86, 37.98, 44.81, 47.77,
49.24, 54.47, 71.06, 123.86, 130.07, 130.64, 140.18, 142.03, 147.45,
209.05. Anal. Calcd. for C26H33NO4: C 73.73, H 7.85, N 3.31; Found:
C 73.62, H 7.91, N 3.19.
To a solution of condensation products (5 mmol) (2a–h) in ace-
tone (20 ml), Jones reagent was added drop wise. The mixture was
stirred at 0 ◦C until the reaction was completed (monitored by TLC).
The reaction was quenched with ethanol (3–5 ml) and concentrated
in reduced pressure. The crude product was dissolved in CH2Cl2,
the organic layer was washed with brine and dried over anhydrous
Na2SO4, filtered. The filtrate was concentrated in reduced pressure
to afford a crude product. The products were purified by flash col-

H. Guo et al. / Steroids 76 (2011) 709–723
711
Scheme 1. Preparation of target compounds (12a–h, 13a–h, 14a–h, 15a–h).
umn chromatography to yield compound 3a–h. The Jones reagent
was prepared according to previous study [21–23].
(150 MHz, CDCl3): ı 11.50, 14.48, 20.75, 28.60, 29.23, 30.76, 31.56,
34.61, 35.90, 38.06, 38.38, 44.56, 46.56, 47.55, 49.27, 53.94, 128.96,
131.43, 131.74, 134.05, 135.16, 136.41, 209.27, 211.51. Anal. Calcd.
for C26H31ClO2: C 75.98, H 7.60; Found: C 75.81, H 7.72.
2.3.1. 16-(4-methyl-benzylidene)-5˛-androstan-3,17-dione (3a)
White solid. Yield 95%. m.p.165-167 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.99 (s, 3H, CH3-19), 1.09 (s, 3H, CH3-18), 2.40 (s, 3H,
Ph-CH3), 7.43 (s, 1H, CH), 7.25 (d, 2H, J = 7.96 Hz), 7.46 (d, 2H,
J = 8.20 Hz). 13C NMR (150 MHz, CDCl3): ı 11.50, 14.54, 20.77, 21.49,
28.64, 29.31, 30.76, 31.60, 34.60, 35.89, 38.07, 38.33, 44.58, 46.56,
47.49, 49.38, 53.94, 129.46, 130.37, 132.78, 133.16, 134.94, 209.65,
211.57. Anal. Calcd. for C27H34O2: C 83.03, H 8.77; Found: C 83.31,
H 8.62.
2.3.6. 16-(4-bromo-benzylidene)-5˛-androstan-3,17-dione (3f)
White solid. Yield 98%. m.p. 182–184 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.99 (s, 3H, CH3-19), 1.09 (s, 3H, CH3-18), 7.37 (s, 1H,
CH), 7.39 (d, 2H, J = 8.48 Hz), 7.40 (d, 2H, J = 8.44 Hz). 13C NMR
(150 MHz, CDCl3): ı 11.51, 14.48, 20.74, 28.59, 29.26, 30.75, 31.55,
34.60, 35.89, 38.07, 38.33, 44.56, 46.55, 47.57, 49.25, 53.92, 123.52,
131.66, 131.81, 131.93, 134.46, 136.57, 209.30, 211.58. Anal. Calcd.
for C26H31BrO2: C 68.57, H 6.86; Found: C 68.44, H 6.94.
2.3.2. 16-(4-methoxy-benzylidene)-5˛-androstan-3,17-dione
(3b)
2.3.7. 16-(4-fluoro-benzylidene)-5˛-androstan-3,17-dione (3g)
White solid. Yield 90%. m.p. 208–210 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.99 (s, 3H, CH3-19), 1.09 (s, 3H, CH3-18), 7.41 (s, 1H,
CH), 7.29 (d, 2H, J = 8.44 Hz), 7.12 (t, 2H), 7.54 (t, 2H). 13C NMR
(150 MHz, CDCl3): ı 11.48, 14.49, 20.73, 28.59, 29.14, 30.74, 31.55,
34.58, 35.87, 38.05, 38.31, 44.55, 46.54, 47.51, 49.29, 53.91, 115.94,
132.22, 161.76, 164.26, 209.39, 211.56. Anal. Calcd. for C26H31FO2:
C 79.15, H 7.92; Found: C 79.28, H 7.88.
White solid. Yield 97%. m.p. 273–275 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.96 (s, 3H, CH3-19), 1.07 (s, 3H, CH3-18), 3.84 (s, 3H,
OCH3), 7.39 (s, 1H, CH), 6.94 (d, 2H, J = 8.76 Hz), 7.51 (d, 2H,
J = 8.76 Hz). 13C NMR (150 MHz, CDCl3): ı 11.47, 14.56, 20.76, 28.63,
29.22, 30.74, 31.59, 34.57, 35.86, 38.05, 38.30, 44.56, 46.53, 47.39,
49.40, 53.93, 55.34, 114.21, 128.23, 132.07, 132.86, 133.49, 160.46,
209.61, 211.50. Anal. Calcd. for C27H34O3: C 79.76, 8.43; Found: C
79.62, H 8.70.
2.3.8. 16-(furan-2-ylmethylene)-5˛-androstan-3,17-dione (3h)
Yellow solid. Yield 97%. m.p. 197–199 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.95 (s, 3H, CH3-19), 1.08 (s, 3H, CH3-18), 7.21 (s, 1H, CH),
6.52 (q, 1H), 6.66 (d, 1H, J = 3.40 Hz), 7.57 (d, 1H, J = 1.68 Hz). 13C
NMR (150 MHz, CDCl3): ı 11.46, 14.58, 20.75, 28.63, 28.85, 30.73,
31.52, 34.53, 35.86, 38.05, 38.31, 44.56, 46.55, 47.64, 48.89, 53.93,
209.42, 211.54. Anal. Calcd. for C24H30O3: C 78.65, H 8.25; Found:
C 78.77, H 8.30.
2.3.3. 16-(4-nitro-benzylidene)-5˛-androstan-3,17-dione (3c)
Yellow solid. Yield 96%. m.p. 236–238 ◦C; 1H NMR (400 MHz,
CDCl3): ı 1.00 (s, 3H, CH3-19), 1.08 (s, 3H, CH3-18), 7.44 (s, 1H,
CH), 7.67 (d, 2H, J = 8.80 Hz), 8.27 (d, 2H, J = 8.80 Hz). 13C NMR
(150 MHz, CDCl3): ı 11.46, 14.40, 20.68, 28.52, 29.31, 30.69, 31.50,
34.56, 35.86, 38.00, 38.28, 44.49, 46.49, 47.65, 49.02, 53.85, 123.83,
130.14, 130.66, 139.97, 141.94, 147.42, 208.62, 211.36. Anal. Calcd.
for C26H31NO4: C 74.08, H 7.41, N 3.32; Found: C 74.15, H 7.52, N
3.40.
2.4. General procedure for the synthesis of compounds 4a–h,
5a–h, 6a–h, 7a–h
2.3.4. 16-(3,4,5-trimethoxy-benzylidene)-5˛-androstan-3,17-
dione (3d)
White solid. Yield 98%. m.p. 211–213 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.86 (s, 3H, CH3-19), 1.07 (s, 3H, CH3-18), 3.89 (s, 9H),7.36
(s, 1H, CH), 6.77 (s, 2H). 13C NMR (150 MHz, CDCl3): ı 11.45, 14.50,
20.72, 22.68, 29.12, 29.35, 29.69, 30.73, 31.55, 31.91, 34.54, 35.87,
38.03, 38.30, 44.53, 46.53, 47.49, 49.32, 53.89, 56.14, 60.93, 107.65,
131.06, 133.29, 134.99, 139.30, 153.17, 209.34, 211.48. Anal. Calcd.
for C29H38O5: C 74.65, H 8.21; Found: C 74.58, H 8.37.
To a suspension of dione 3a–h (1 mmol) in 20 ml of absolute
methanol was added 0.5 ml of piperidine and 60 mg (1.2 mmol,
1.2 equive.) of NaBH3CN, then the solution was acidified to pH ≈ 6
with acetic acid. The resulting mixture was stirred at 40 ◦C for 8 h.
The methanol was removed under reduced pressure, 20 ml CH2Cl2
was added to the residue, the organic layer was washed with sat-
urated NaHCO3, brine and dried over anhydrous Na2SO4, filtered.
The filtrate was concentrated in vacuum to afford a crude product.
The product was purified by flash column chromatography to yield
compound 4a–h. Compounds 5a–h, 6a–h and 7a–h were obtained
with pyrrolidine, morpholine, N-methylpiperazine by using the
same method that described above [24].
2.3.5. 16-(4-chloro-benzylidene)-5˛-androstan-3,17-dione (3e)
White solid. Yield 98%. m.p. 256–258 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.90 (s, 3H, CH3-19), 1.00 (s, 3H, CH3-18), 7.31 (s, 1H,
CH), 7.29 (d, 2H, J = 8.44 Hz), 7.40 (d, 2H, J = 8.56 Hz). 13C NMR

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H. Guo et al. / Steroids 76 (2011) 709–723
2.4.1. 16-(4-methyl-benzylidene)-3ˇ-piperidino-5˛-androstan-
17-one (4a)
132.06, 132.65, 133.86, 160.38, 210.21. Anal. Calcd. for C31H43NO2:
C 80.65, H 9.39, N 3.03; Found C 80.51, H 9.18, N 3.11.
White solid. Yield 45%. m.p 165–167 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.81 (s, 3H, CH3-19), 0.97 (s, 3H, CH3-18), 2.40 (s, 3H, Ph-
CH3), 2.53 (s, 4H, NCH2), 7.42 (s, 1H, CH), 7.15 (d, 2H, J = 7.96 Hz),
7.27 (d, 2H, J = 8.20 Hz). 13C NMR (150 MHz, CDCl3): ı 12.40, 14.57,
20.43, 24.10, 24.77, 26.38, 28.77, 29.19, 30.68, 31.22, 31.66, 34.68,
36.44, 37.89, 46.11, 47.59, 49.52, 50.42, 54.60, 55.31, 64.79, 114.19,
128.46, 132.08, 132.47, 134.96, 209.76. Anal. Calcd. for C32H45NO:
C 83.61, H 9.87, N 3.05; Found C 83.55, H 9.91, N 3.12.
2.4.7. 16-(4-methoxy-benzylidene)-3ˇ-morpholino-5˛-
androstan-17-one (6b)
White solid. Yield 41%. m.p. 214–216 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.84 (s, 3H, CH3-19), 0.94 (s, 3H, CH3-18), 2.57 (s, 4H,
NCH2), 3.72–3.75 (m, 4H, OCH2), 3.84 (s, 3H, OCH3), 7.38 (s, 1H,
CH), 6.92 (d, 2H, J = 8.76 Hz), 7.49 (d, 2H, J = 8.80 Hz). 13C NMR
(150 MHz, CDCl3): ı 12.39, 14.58, 20.48, 24.49, 28.74, 29.27, 31.09,
31.18, 31.69, 34.71, 36.12, 37.62, 45.73, 47.54, 49.64, 50.05, 54.64,
55.35, 64.05, 67.34, 114.17, 128.36, 132.06, 132.71, 133.79, 160.40,
210.13. Anal. Calcd. for C31H43NO3: C 77.95, H 9.07, N 2.93; Found
C 77.88, H 9.16, N 2.88.
2.4.2. 16-(4-methyl-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17-one (5a)
White solid. Yield 48%. m.p 206–208 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.87 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.40 (s, 3H, Ph-
CH3), 2.57 (br, 4H, NCH2), 7.42 (s, 1H, CH), 7.22 (d, 2H, J = 7.96 Hz),
7.44 (d, 2H, J = 8.08 Hz). 13C NMR (150 MHz, CDCl3): ı 12.07, 14.52,
20.43, 21.46, 23.26, 24.16, 28.22, 29.27, 30.67, 30.91, 31.57, 34.55,
35.75, 36.75, 45.18, 47.53, 49.49, 50.66, 54.22, 63.96, 129.44, 130.35,
132.76, 133.14, 135.01, 139.63, 176.31, 209.94. Anal. Calcd. for
2.4.8. 16-(4-methoxy-benzylidene)-3ˇ-(4-methylpiperazinyl)-
5˛-androstan-17-one (7b)
White solid. Yield 44%. m.p. 239–241 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.83 (s, 3H, CH3-19), 0.93 (s, 3H, CH3-18), 2.28 (s, 3H,
N-CH3), 2.40–2.61 (m, 8H, NCH2), 3.84 (s, 3H, OCH3), 7.38 (s, 1H,
CH), 6.92 (d, 2H, J = 8.68 Hz), 7.49 (d, 2H, J = 8.68 Hz). 13C NMR
(150 MHz, CDCl3): ı 12.41, 14.58, 20.47, 24.55, 28.73, 29.27, 31.19,
31.23, 31.69, 34.71, 36.11, 37.73, 45.85, 46.03, 47.54, 49.28, 49.63,
54.65, 55.35, 55.49, 63.75, 114.16, 128.36, 132.05, 132.67, 132.81,
160.38, 210.14. Anal. Calcd. for C32H46N2O2: C 78.32, H 9.45, N 5.71;
Found C 78.21, H 9.58, N 5.66.
C31H43NO: C 83.54, H 9.72, N 3.14; Found C 83.49, H 9.88, N 3.22.
2.4.3. 16-(4-methyl-benzylidene)- 3ˇ-morpholino-
5˛-androstan-17-one (6a)
White solid. Yield 42%. m.p. 234–236 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.76 (s, 3H, CH3-19), 0.87 (s, 3H, CH3-18), 2.31 (s, 3H,
Ph-CH3), 2.49 (s, 4H, NCH2), 3.63–3.65 (m, 4H, OCH2), 7.33 (s, 1H,
CH), 7.13 (d, 2H, J = 7.96 Hz), 7.35 (d, 2H, J = 8.08 Hz). 13C NMR
(150 MHz, CDCl3): ı 12.18, 14.52, 20.44, 21.46, 21.98, 22.38, 28.40,
28.88, 29.27, 30.92, 31.57, 34.59, 35.87, 37.01, 45.34, 47.54, 48.60,
49.46, 54.25, 64.70, 64.99, 129.44, 130.36, 132.78, 133.16, 135.02,
139.61, 176.08, 209.98. Anal. Calcd. for C31H43NO2: C 80.65, H 9.39,
N 3.03; Found C 80.51, H 9.47, N 3.22.
2.4.9. 16-(4-nitro-benzylidene)-3ˇ-piperidino-5˛-
androstan-17-one (4c)
Yellow solid. Yield 44%. m.p. 243–245 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.85 (s, 3H, CH3-19), 0.98 (s, 3H, CH3-18), 2.54 (s, 4H,
NCH2), 7.45 (s, 1H, CH), 7.67 (d, 2H, J = 8.92 Hz), 8.26 (d, 2H,
J = 8.80 Hz). 13C NMR (150 MHz, CDCl3): ı 12.42, 14.42, 20.41, 24.08,
24.82, 26.36, 28.64, 29.25, 29.39, 30.62, 31.17, 31.61, 34.72, 36.19,
37.90, 46.03, 47.79, 49.26, 50.35, 53.77, 54.59, 64.43, 123.85, 129.98,
130.62, 140.24, 142.06, 147.42, 209.09. Anal. Calcd. for C31H42N2O3:
C 75.88, H 8.63, N 5.71; Found C 75.97, H 8.58, N 5.62.
2.4.4. 16-(4-methyl-benzylidene)-3ˇ-(4-Methylpiperazinyl)-5˛-
androstan-17-one (7a)
White solid. Yield 43%. m.p. 256–258 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.76 (s, 3H, CH3-19), 0.86 (s, 3H, CH3-18), 2.31 (s, 3H,
Ph-CH3), 2.21 (s, 3H, N-CH3), 2.39–2.53 (m, 8H, NCH2), 7.32 (s, 1H,
CH), 7.13 (d, 2H, J = 7.96 Hz), 7.35 (d, 2H, J = 8.08 Hz). 13C NMR
(150 MHz, CDCl3): ı 12.42, 14.54, 20.47, 21.47, 24.58, 28.73, 29.34,
31.19, 31.27, 31.69, 34.73, 36.12, 37.74, 45.86, 46.05, 47.60, 49.32,
49.59, 54.65, 55.53, 63.77, 129.42, 130.34, 132.88, 132.91, 135.25,
139.49, 210.08. Anal. Calcd. for C32H46N2O: C 80.96, H 9.77, N 5.90;
Found C 81.22, H 9.88, N 5.80.
2.4.10. 16-(4-nitro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17-one (5c)
Yellow solid. Yield 41%. m.p. 225–227 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.87 (s, 3H, CH3-19), 0.98 (s, 3H, CH3-18), 2.59 (br, 4H,
NCH2), 7.45 (s, 1H, CH), 7.67 (d, 2H, J = 8.80 Hz), 8.26 (d, 2H,
J = 8.84 Hz). 13C NMR (150 MHz, CDCl3): ı 12.35, 14.43, 20.35, 23.17,
27.99, 28.57, 29.40, 31.19, 31.62, 34.74, 36.02, 37.34, 45.25, 47.80,
49.30, 51.85, 54.61, 64.26, 123.85, 129.95, 130.62, 140.26, 142.06,
147.42, 209.08. Anal. Calcd. for C30H40N2O3: C 75.59, H 8.46, N 5.88;
Found C 75.71, H 8.31, N 5.97.
2.4.5. 16-(4-methoxy-benzylidene)-3ˇ-piperidino-5˛-
androstan-17-one (4b)
White solid. Yield 45%. m.p. 215–217 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.84 (s, 3H, CH3-19), 0.98 (s, 3H, CH3-18), 2.53 (s, 4H,
NCH2), 3.86 (s, 3H, OCH3), 7.40 (s, 1H, CH), 6.94 (d, 2H, J = 8.72 Hz),
7.51 (d, 2H, J = 8.72 Hz). 13C NMR (150 MHz, CDCl3): ı 12.44, 14.59,
20.49, 24.12, 24.87, 26.43, 28.77, 29.29, 30.73, 31.22, 31.71, 34.73,
36.21, 37.94, 46.09, 47.57, 49.66, 50.37, 54.69, 55.36, 64.48, 114.16,
128.38, 132.05, 132.66, 133.85, 160.37, 209.83. Anal. Calcd. for
2.4.11. 16-(4-nitro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17-one (6c)
Yellow solid. Yield 44%. m.p. 241–243 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.88 (s, 3H, CH3-19), 1.00 (s, 3H, CH3-18), 2.59 (s, 4H,
NCH2), 3.73–3.76 (s, 4H, OCH2), 7.45 (s, 1H, CH), 7.67 (d, 2H,
J = 8.76 Hz), 8.26 (d, 2H, J = 8.80 Hz). 13C NMR (150 MHz, CDCl3): ı
12.38, 14.43, 20.41, 24.47, 28.62, 29.38, 31.04, 31.14, 31.60, 34.72,
36.12, 37.61, 45.69, 47.79, 49.26, 50.04, 54.56, 64.00, 67.32, 123.86,
130.04, 130.63, 140.19, 142.04, 147.44, 209.07. Anal. Calcd. for
C32H45NO2: C 80.79, H 9.53, N 2.94; Found C 80.91, H 9.66, N 2.81.
2.4.6. 16-(4-methoxy-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17-one (5b)
C30H40N2O4: C 73.14, H 8.18, N 5.69; Found C 73.22, H 8.26, N 5.55.
White solid. Yield 47%. m.p. 224–226 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.86 (s, 3H, CH3-19), 0.93 (s, 3H, CH3-18), 2.57 (br, 4H,
NCH2), 3.85 (s, 3H, OCH3), 7.38 (s, 1H, CH), 6.92 (d, 2H, J = 8.80 Hz),
7.49 (d, 2H, J = 8.80 Hz). 13C NMR (150 MHz, CDCl3): ı 12.36, 14.58,
20.43, 23.17, 27.98, 28.68, 29.29, 31.23, 31.71, 34.73, 34.76, 36.02,
37.35, 45.29, 47.56, 49.68, 51.83, 54.69, 55.35, 64.29, 114.16, 128.39,
2.4.12. 16-(4-nitro-benzylidene)-3ˇ-(4-Methylpiperazinyl)-5˛-
androstan-17-one (7c)
Yellow solid. Yield 44%. m.p. 259–261 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.85 (s, 3H, CH3-19), 0.98 (s, 3H, CH3-18), 2.30 (s, 3H,
N-CH3), 2.42–2.62 (m, 8H, NCH2), 7.45 (s, 1H, CH), 7.67 (d, 2H,
J = 8.76 Hz), 8.26 (d, 2H, J = 8.76 Hz). 13C NMR (150 MHz, CDCl3): ı

H. Guo et al. / Steroids 76 (2011) 709–723
713
12.40, 14.43, 20.40, 24.53, 28.62, 29.38, 31.15, 31.19, 31.60, 34.72,
36.11, 37.72, 45.81, 46.01, 47.79, 49.28, 54.57, 55.47, 63.72, 123.85,
130.00, 130.62, 140.22, 142.05, 147.43, 209.07. Anal. Calcd. for
NCH2), 7.39 (s, 1H, CH), 7.40 (d, 2H, J = 8.56 Hz), 7.49 (d, 2H,
J = 8.48 Hz). 13C NMR (150 MHz, CDCl3): ı 12.35, 14.48, 20.38, 23.18,
23.66, 28.01, 28.63, 29.26, 31.20, 31.66, 34.72, 34.77, 36.01, 37.34,
45.26, 47.64, 49.48, 51.85, 54.63, 64.27, 128.91, 131.42, 134.14,
135.02, 136.74, 209.63. Anal. Calcd. for C30H40ClNO: C 77.31, H 8.65,
N 3.01; Found C 77.44, H 8.78, N 2.99.
C31H43N3O3: C 73.63, H 8.57, N 8.31; Found C 73.81, H 8.42, N 8.26.
2.4.13. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-piperidino-5˛-
androstan-17-one (4d)
White solid. Yield 47%. m.p. 197–199 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.82 (s, 3H, CH3-19), 0.95 (s, 3H, CH3-18), 2.51 (s, 4H,
NCH2), 3.89 (s, 9H, OCH3), 7.34 (s, 1H, CH), 6.77 (s, 2H). 13C NMR
(150 MHz, CDCl3): ı 12.41, 14.53, 20.45, 24.10, 24.86, 26.41, 28.72,
29.18, 30.71, 31.20, 31.67, 34.70, 36.20, 37.91, 46.06, 47.64, 49.55,
50.36, 54.63, 56.15, 60.97, 64.45, 107.62, 131.19, 133.02, 135.37,
139.21, 153.17, 209.83. Anal. Calcd. for C34H49NO4: C 76.22, H 9.22,
N 2.61; Found C 76.41, H 9.31, N 2.48.
2.4.19. 16-(4-chloro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17-one (6e)
White solid. Yield 49%. m.p. 220–222 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.86 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.57 (s,
4H, NCH2), 3.74 (s, 4H, OCH2), 7.39 (s, 1H, CH), 7.40 (d, 2H,
J = 8.56 Hz), 7.49 (d, 2H, J = 8.52 Hz). 13C NMR (150 MHz, CDCl3): ı
12.39, 14.49, 20.44, 24.49, 28.68, 29.25, 31.08, 31.16, 31.63, 34.70,
36.11, 37.61, 45.71, 47.65, 49.45, 50.05, 54.59, 64.03, 67.34, 128.93,
131.42, 131.51, 134.12, 135.05, 136.68, 209.67. Anal. Calcd. for
2.4.14. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17-one (5d)
C30H40ClNO2: C 74.74, H 8.36, N 2.91; Found C 74.68, H 8.43, 21,
N 2.88.
White solid. Yield 48%. m.p. 162–164 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.85 (s, 3H, CH3-19), 0.95 (s, 3H, CH3-18), 2.56 (br, 4H,
NCH2), 3.89 (s, 9H, OCH3), 7.34 (s, 1H, CH), 6.77 (s, 2H). 13C NMR
(150 MHz, CDCl3): ı 10.73, 18.98, 19.61, 22.82, 27.13, 29.28, 29.47,
30.07, 33.21, 34.37, 34.86, 36.00, 41.35, 44.02, 47.23, 48.41, 52.71,
54.46, 59.29, 62.45, 65.71, 82.93, 103.93, 122.00, 131.76, 135.22,
138.82, 151.34, 169.62. Anal. Calcd. for C33H47NO4: C 75.97, H 9.08,
N 2.68; Found C 75.88, H 9.12, N 2.79.
2.4.20. 16-(4-chloro-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17-one (7e)
White solid. Yield 47%. m.p. 265–267 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.76 (s, 3H, CH3-19), 0.87 (s, 3H, CH3-18), 2.21 (s, 3H,
N-CH3), 2.39–2.55 (m, 8H, NCH2), 7.29 (s, 1H, CH), 7.31 (d, 2H,
J = 8.60 Hz), 7.39 (d, 2H, J = 8.48 Hz). 13C NMR (150 MHz, CDCl3): ı
12.41, 14.49, 20.44, 24.53, 28.68, 29.26, 31.17, 31.22, 31.65, 34.72,
36.12, 37.73, 45.84, 46.03, 47.66, 49.31, 49.47, 54.62, 55.47, 63.79,
128.93, 131.42, 131.48, 134.14, 135.04, 136.72, 209.68. Anal. Calcd.
for C31H43ClN2O: C 75.20, H 8.75, N 5.66; Found C 75.11, H 8.99, N
5.49.
2.4.15. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-morpholino-5˛-
androstan-17-one (6d)
White solid. Yield 48%. m.p. 168–170 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.86 (s, 3H, CH3-19), 0.95 (s, 3H, CH3-18), 2.56 (s, 4H,
NCH2), 3.73–3.75 (m, 4H, OCH2), 3.89 (s, 9H, OCH3), 7.35 (s, 1H,
CH), 6.77 (s, 2H). 13C NMR (150 MHz, CDCl3): ı 12.37, 14.52, 20.43,
24.50, 28.68, 29.92, 31.10, 31.16, 31.64, 34.66, 35.64, 36.11, 37.60,
45.70,47.53, 49.52, 50.05, 53.78, 54.57, 56.14, 60.97, 67.34, 107.58,
131.16, 133.04, 133.27, 135.04, 135.31, 139.19, 153.16, 209.75.
Anal. Calcd. for C33H47NO5: C 73.71, H 8.81, N 2.60; Found C 73.52,
H 8.93, N 2.46.
2.4.21. 16-(4-bromo-benzylidene)-3ˇ-piperidino-5˛-
androstan-17-one (4f)
White solid. Yield 45%. m.p. 207–209 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.86 (s, 3H, CH3-19), 0.97 (s, 3H, CH3-18), 2.55 (s, 4H,
NCH2), 7.37 (s, 1H, CH), 7.40 (d, 2H, J = 8.52 Hz), 7.54 (d, 2H,
J = 8.61 Hz). 13C NMR (150 MHz, CDCl3): ı 12.34, 14.49, 20.55, 22.66,
28.38, 29.29, 29.71, 31.12, 31.44, 34.71, 35.74, 36.86, 38.04, 44.83,
45.99, 47.66, 49.44, 51.63, 54.50, 71.14, 123.44, 131.65, 131.19,
134.54, 136.84, 209.66. Anal. Calcd. for C31H42BrNO: C 70.98, H 8.07,
N 2.67; Found C 70.77, H 7.98, N 2.71.
2.4.16. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-(4-
methylpiperazinyl)-5˛-androstan-17-one (7d)
White solid. Yield 49%. m.p. 185–187 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.85 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.31 (s, 3H, N-
CH3), 2.48–2.65 (m, 8H, NCH2), 3.89 (s, 9H, OCH3), 7.35 (s, 1H, CH),
6.78 (s, 2H). 13C NMR (150 MHz, CDCl3): ı 12.36,14.51, 20.41, 24.44,
28.65, 29.14, 31.15, 31.62, 34.64, 36.07, 37.66, 45.78, 47.59, 49.14,
49.50, 54.55, 55.31, 56.12, 60.93, 63.70, 107.57, 131.15, 133.00,
135.30, 139.17, 153.13, 209.71. Anal. Calcd. for C34H50N2O4: C
74.14, H 9.15, N 5.09; Found C 74.26, H 9.22, N 5.01.
2.4.22. 16-(4-bromo-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17-one (5f)
White solid. Yield 41%. m.p. 242–244 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.87 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.59 (br, 4H,
NCH2), 7.36 (s, 1H, CH), 7.40 (d, 2H, J = 8.52 Hz), 7.54 (d, 2H,
J = 8.61 Hz). 13C NMR (150 MHz, CDCl3): ı 12.36, 14.49, 20.39, 23.18,
27.91, 28.62, 29.29, 31.20, 31.66, 34.66, 34.72, 36.02, 37.33, 45.26,
47.68, 49.48, 51.83, 54.62, 64.30, 123.38, 131.50, 131.63, 131.87,
134.56, 136.91, 209.71. Anal. Calcd. for C30H40BrNO: C 70.58, H 7.90,
N 2.74; Found C 70.44, H 7.81, N 2.88.
2.4.17. 16-(4-chloro-benzylidene)-3ˇ-piperidino-5˛-
androstan-17-one (4e)
White solid. Yield 49%. m.p. 163–165 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.91 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.51 (s,
4H, NCH2), 7.39 (s, 1H, CH), 7.40 (d, 2H, J = 8.44 Hz), 7.48
(d, 2H, J = 8.52 Hz). 13C NMR (150 MHz, CDCl3): ı 12.33, 14.49,
20.45, 22.13, 22.75, 23.03, 28.20, 28.79, 29.17, 30.85, 31.50, 34.55,
35.84, 36.99, 45.43, 47.56, 49.31, 49.93, 54.12, 65.45, 128.95,
131.43, 131.72, 134.03, 135.14, 136.43, 209.43. Anal. Calcd. for
2.4.23. 16-(4-bromo-benzylidene)-3ˇ-morpholino-5˛-
androstan-17-one (6f)
White solid. Yield 47%. m.p. 247–249 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.67 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.58 (s, 4H,
NCH2), 3.73–3.75 (m, 4H, OCH2), 7.36 (s, 1H, CH), 7.39 (d, 2H,
J = 8.52 Hz), 7.54 (d, 2H, J = 8.48 Hz). 13C NMR (150 MHz, CDCl3): ı
12.38, 14.49, 20.34, 24.50, 28.66, 29.29, 31.02, 31.11, 31.61, 34.73,
36.11, 37.58, 45.73, 47.66, 49.35, 50.15, 54.49, 64.00, 67.27, 128.88,
131.42, 131.49, 134.11, 135.03, 136.71, 209.77. Anal. Calcd. for
C31H42ClNO: C 77.55, H 8.82, N 2.92; Found C 77.71, H 8.76, N
2.89.
2.4.18. 16-(4-chloro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17-one (5e)
White solid. Yield 49%. m.p. 232–234 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.87 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.57 (br, 4H,
C30H40BrNO2: C 68.43, H 7.66, N 2.66; Found C 68.51, H 7.59, N
2.71.

714
H. Guo et al. / Steroids 76 (2011) 709–723
2.4.24. 16-(4-bromo-benzylidene)-3ˇ-(4-methylpiperazinyl)-
5˛-androstan-17-one (7f)
144.62, 152.32, 209.97. Anal. Calcd. for C29H41NO2: C 79.95, H 9.49,
N 3.22; Found C 79.80, H 9.61, N 3.08.
White solid. Yield 40%. m.p. 290–292 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.85 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.30 (s, 3H,
N-CH3), 2.49–2.62 (m, 8H, NCH2), 7.36 (s, 1H, CH), 7.39 (d, 2H,
J = 8.32 Hz), 7.54 (d, 2H, J = 8.24 Hz). 13C NMR (150 MHz, CDCl3): ı
12.38, 14.55, 20.51, 24.49, 28.73, 29.22, 31.27, 31.19, 31.58, 34.66,
36.33, 37.73, 45.91, 46.13, 47.72, 49.41, 49.48, 54.64, 55.43, 63.81,
128.98, 131.44, 131.54, 134.26, 135.10, 136.77, 209.71. Anal. Calcd.
for C31H43BrN2O: C 69.00, H 8.03, N 5.19; Found C 69.13, H 8.01, N
5.27.
2.4.30. 16-(furan-2-ylmethylene)-3ˇ-pyrrolidino-5˛-
androstan-17-one (5h)
White solid. Yield 48%. m.p. 228–230 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.86 (s, 3H, CH3-19), 0.93 (s, 3H, CH3-18), 2.58 (br, 4H,
NCH2), 7.21 (s, 1H, CH), 6.52 (q, 1H), 6.65 (d, 2H, J = 3.52 Hz), 7.57
(s, 1H). 13C NMR (150 MHz, CDCl3): ı 12.35, 14.61, 20.42, 23.17,
27.95, 28.68, 28.91, 31.22, 31.64, 34.70, 34.73, 36.01, 37.35, 45.30,
47.79, 49.15, 51.82, 54.68, 64.30, 112.29, 115.40, 119.41, 133.78,
144.60, 152.30, 209.93. Anal. Calcd. for C28H39NO2: C 79.76, H 9.32,
N 3.32; Found C 79.52, H 9.44, N 3.19.
2.4.25. 16-(4-fluoro-benzylidene)-3ˇ-piperidino-5˛-
androstan-17-one (4g)
White solid. Yield 43%. m.p. 228–230 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.84 (s, 3H, CH3-19), 0.95 (s, 3H, CH3-18), 2.52 (s, 4H,
NCH2), 7.39 (s, 1H, CH), 7.11 (t, 2H), 7.54 (t, 2H). 13C NMR
(150 MHz, CDCl3): ı 12.41, 14.48, 20.44, 24.14, 24.85, 26.42, 28.71,
29.16, 30.69, 31.18, 31.65, 34.70, 36.18, 37.91, 46.05, 47.59, 49.50,
50.37, 54.63, 64.47, 115.66, 115.88, 131.57, 132.08, 132.17, 135.73,
135.75, 161.70, 164.19, 209.67. Anal. Calcd. for C31H42FNO: C 80.30,
H 9.13, N 3.02; Found C 80.11, H 9.08, N 2.88.
2.4.31. 16-(furan-2-ylmethylene)-3ˇ-morpholino-5˛-
androstan-17-one (6h)
White solid. Yield 42%. m.p.232–234 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.85 (s, 3H, CH3-19), 0.92 (s, 3H, CH3-18), 2.58 (s, 4H,
NCH2), 3.73–3.75 (m, 4H, OCH2), 7.20 (s, 1H, CH), 6.52 (q, 1H), 6.65
(d, 2H, J = 3.40 Hz), 7.57 (s, 1H). 13C NMR (150 MHz, CDCl3): ı 12.39,
14.61, 20.48, 24.47, 28.74, 28.90, 31.08, 31.17, 31.62, 34.68, 36.11,
37.62, 45.74, 47.77, 49.11, 50.04, 54.64, 64.08, 67.30, 112.31, 115.46,
119.45, 133.70, 144.64, 152.27, 209.87. Anal. Calcd. for C28H39NO3:
C 76.85, H 8.98, N 3.20; Found C 76.62, H 8.76, N 3.39.
2.4.26. 16-(4-fluoro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17-one (5g)
2.4.32. 16-(furan-2-ylmethylene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17-one (7h)
White solid. Yield 46%. m.p. 265–267 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.78 (s, 3H, CH3-19), 0.87 (s, 3H, CH3-18), 2.49 (br,
4H, NCH2), 7.31 (s, 1H, CH), 7.02 (t, 2H), 7.45 (t, 2H). 13C NMR
(150 MHz, CDCl3): ı 12.35, 14.51, 20.39, 23.17, 27.91, 28.63, 29.20,
31.21, 31.66, 34.68, 34.73, 36.02, 37.33, 45.27, 47.64, 49.55, 51.83,
54.64, 64.30, 115.69, 115.91, 131.63, 131.91, 132.11, 132.19, 135.80,
161.74, 164.23, 209.87. Anal. Calcd. for C30H40FNO: C 80.13, H 8.97,
N 3.12; Found C 80.22, H 8.77, N 3.25.
White solid. Yield 49%. m.p. 218–220 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.84 (s, 3H, CH3-19), 0.92 (s, 3H, CH3-18), 2.30 (s, 3H,
N-CH3), 2.46–2.62 (m, 8H, NCH2), 7.21 (s, 1H, CH), 6.52 (q, 1H),
6.65 (d, 2H, J = 3.42 Hz), 7.57 (s, 1H). 13C NMR (150 MHz, CDCl3): ı
12.41, 14.61, 20.47, 24.56, 28.74, 28.90, 31.19, 31.27, 31.62, 34.69,
36.11, 37.74, 45.87, 46.05, 47.79, 49.11, 49.31, 54.66, 55.51, 63.77,
112.30, 115.44, 119.44, 133.74, 144.62, 152.28, 209.95. Anal. Calcd.
for C29H42N2O2: C 77.29, H 9.39, N 6.22; Found C 77.41, H 9.51, N
6.08.
2.4.27. 16-(4-fluoro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17-one (6g)
White solid. Yield 45%. m.p. 236–238 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.86 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.58 (s, 4H,
NCH2), 3.72–3.74 (m, 4H, OCH2), 7.40 (s, 1H, CH), 7.12 (t, 2H),
7.54 (t, 2H).13C NMR (150 MHz, CDCl3): ı 12.39, 14.51, 20.45, 24.51,
28.70, 29.18, 31.09, 31.17, 31.64, 34.71, 36.12, 37.62, 45.72, 47.63,
49.51, 50.06, 54.60, 64.04, 67.35, 115.70, 115.92, 131.68, 131.86,
131.89, 132.11, 132.19, 161.74, 164.24, 209.80. Anal. Calcd. for
C30H40FNO2: C 77.38, H 8.66, N 3.01; Found C 77.47, H 8.51, N 2.92.
2.5. General procedure for the synthesis of compounds
8a–h,9a–h, 10a–h, 11a–h
To the suspension of compounds 4a–h, 5a–h, 6a–h, 7a–h
(1 mmol) in 20 ml of absolute methanol at 0 ◦C, sodium borohy-
dride (1.2 equiv.) was added in small amounts over a period of 3 h.
The methanol was removed under reduced pressure, 20 ml CH2Cl2
was added to the residue, the organic layer was washed with sat-
urated NaHCO3, brine and dried over anhydrous Na2SO4, filtered.
The filtrate was concentrated in vacuum to afford a crude product.
The product was purified by flash column chromatography to yield
compound 8a–h, 9a–h, 10a–h, 11a–h.
2.4.28. 16-(4-fluoro-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17-one (7g)
White solid. Yield 48%. m.p. 245–247 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.85 (s, 3H, CH3-19), 0.96 (s, 3H, CH3-18), 2.30 (s, 3H, N-
CH3), 2.49–2.63 (m, 8H, NCH2), 7.39 (s, 1H, CH), 7.11 (t, 2H), 7.54 (t,
2H). 13C NMR (150 MHz, CDCl3): ı 12.40, 14.49, 20.43, 24.55, 28.68,
29.17, 31.17, 31.22, 31.64, 34.71, 36.10, 37.72, 45.83, 46.00, 47.61,
49.26, 49.50, 54.61, 55.46, 63.74, 115.68, 115.89, 131.61, 131.86,
131.89, 132.09, 132.17, 135.75, 161.72, 164.21, 209.74. Anal. Calcd.
for C31H43FN2O: C 77.78, H 9.05, N 5.85; Found C 77.91, H 8.99, N
5.72.
2.5.1. 16-(4-methyl-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-ol (8a)
White solid. Yield 42%. m.p. 116–118 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.60 (s, 3H, CH3-19), 0.73 (s, 3H, CH3-18), 2.40 (s, 3H,
Ph-CH3), 2.59 (s, 4H, NCH2), 3.90 (s, 1H, C17-␣H), 6.40 (s, 1H, CH),
7.07 (d, 2H, J = 7.92 Hz), 7.20 (d, 2H, J = 7.96 Hz). 13C NMR (150 MHz,
CDCl3): ı 11.26, 12.27, 20.79, 21.14, 21.81, 22.75, 22.96, 23.11,
28.14, 28.44, 30.83, 31.47, 34.86, 35.79, 36.38, 37.05, 42.91, 45.38,
48.51, 48.98, 54.31, 64.44, 84.60, 122.53, 128.01, 129.00, 135.24,
135.82, 145.06, 176.90. Anal. Calcd. for C32H47NO: C 83.24, H 10.26,
N 3.03; Found C 83.42, H 10.19, N 2.99.
2.4.29. 16-(furan-2-ylmethylene)-3ˇ-piperidino-5˛-
androstan-17-one (4h)
White solid. Yield 49%. m.p. 198–200 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.84 (s, 3H, CH3-19), 0.92 (s, 3H, CH3-18), 2.53 (s, 4H,
NCH2), 7.20 (s, 1H, CH), 6.51 (q, 1H), 6.64 (d, 2H, J = 3.44 Hz), 7.56 (s,
1H). 13C NMR (150 MHz, CDCl3): ı 12.44, 14.62, 20.50, 24.17, 24.90,
26.49, 28.78, 28.92, 30.77, 31.23, 31.65, 34.72, 36.22, 37.96, 46.13,
47.80, 49.15, 50.39, 54.71, 64.51, 112.30, 115.42, 119.43, 133.81,
2.5.2. 16-(4-methyl-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-ol (9a)
White solid. Yield 42%. m.p. 202–204 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.84 (s, 3H, CH3-18), 2.35 (s, 3H,

H. Guo et al. / Steroids 76 (2011) 709–723
715
Ph-CH3), 2.59 (br, 4H, NCH2), 3.90 (d, 1H, C17-␣H, J = 8.72 Hz), 6.49
(s, 1H, CH), 7.16 (d, 2H, J = 7.96 Hz), 7.30 (d, 2H, J = 8.31 Hz). 13C
NMR (150 MHz, CDCl3): ı 11.17, 12.38, 20.73, 21.17, 23.19, 27.83,
28.78, 30.96, 31.84, 34.59, 35.04, 35.94, 36.45, 37.43, 43.02, 45.34,
48.59, 51.80, 54.71, 64.40, 85.00, 122.61, 128.13, 129.04, 135.12,
35.99, 145.34. Anal. Calcd. for C31H45NO: C 83.17, H 10.13, N 3.13;
Found C 83.27, H 10.25, N 3.01.
129.39, 130.78, 143.84, 158.01. Anal. Calcd. for C31H45NO3: C 77.62,
H 9.46, N 2.92; Found C 77.43, H 9.59, N 2.71.
2.5.8. 16-(4-methoxy-benzylidene)-3ˇ-(4-methylpiperazinyl)-
5˛-androstan-17ˇ-ol (11b)
White solid. Yield 44%. m.p. 147–149 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.67 (s, 3H, CH3-19), 0.80 (s, 3H, CH3-18), 2.29 (s, 3H,
N-CH3), 2.45–2.62 (m, 8H, NCH2), 3.83 (s, 3H, OCH3), 4.02 (s, 1H,
C
17-␣H), 6.44 (d, 1H, CH, J = 2.12 Hz), 6.88 (d, 2H, J = 8.76 Hz), 7.33
2.5.3. 16-(4-methyl-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-ol (10a)
(d, 2H, J = 8.72 Hz). 13C NMR (150 MHz, CDCl3): ı 11.20, 11.38, 12.42,
20.78, 24.52, 28.84, 30.87, 31.25, 31.81, 35.03, 36.02, 36.45, 37.80,
43.01, 45.87, 46.02, 46.11, 48.58, 49.22, 54.67, 55.24, 55.45, 63.81,
84.90, 113.75, 122.13, 129.38, 130.83, 143.91, 157.97. Anal. Calcd.
for C32H48N2O2: C 78.00, H 9.82, N 5.69; Found C 77.82, H 9.93, N
5.43.
White solid. Yield 47%. m.p. 196–198 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.70 (s, 3H, CH3-19), 0.83 (s, 3H, CH3-18), 2.36 (s, 3H,
Ph-CH3), 2.58 (s, 4H, NCH2), 3.72–3.76 (s, 4H, OCH2), 4.04 (d, 1H,
C17-␣H, J = 8.12 Hz), 6.49 (s, 1H, CH), 7.16 (d, 2H, J = 7.96 Hz), 7.30
(d, 2H, J = 8.33 Hz). 13C NMR (150 MHz, CDCl3): ı 11.19, 12.26, 20.77,
21.16, 22.20, 22.85, 28.62, 29.37, 30.90, 31.62, 34.95, 35.86, 36.33,
37.26, 42.99, 45.49, 48.47, 48.82, 54.41, 64.45, 65.54, 84.93, 122.70,
128.13, 129.06, 135.07, 136.05, 145.04. Anal. Calcd. for C31H45NO2:
C 80.30, H 9.78, N 3.02; Found C 80.12, H 9.69, N 3.11.
2.5.9. 16-(4-nitro-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-ol (8c)
Yellow solid. Yield 47%. m.p. 227–229 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.71 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.54 (s, 4H,
NCH2), 4.09 (s, 1H, C17-␣H), 6.62 (d, 1H, CH, J = 2.32 Hz), 7.52 (d,
2H, J = 8.88 Hz), 8.21 (d, 2H, J = 8.84 Hz). 13C NMR (150 MHz, CDCl3):
ı 11.23, 12.43, 20.71, 24.05, 24.79, 26.29, 28.76, 30.52, 31.23, 31.81,
34.98, 36.12, 37.97, 43.16, 46.09, 48.48, 50.31, 54.65, 64.50, 85.05,
121.42, 123.75, 128.54, 144.51, 145.65, 152.09. Anal. Calcd. for
2.5.4. 16-(4-methyl-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-ol (11a)
White solid. Yield 41%. m.p. 205–207 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.70 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.39 (s, 3H,
Ph-CH3), 2.43–2.63 (m, 8H, NCH2), 3.92 (d, 1H, C17-␣H, J = 8.72 Hz),
C31H44N2O3: C 75.57, H 9.00, N 5.69; Found C 75.64, H 8.93, N 5.80.
6.49 (s, 1H, CH), 7.17 (d, 2H, J = 7.96 Hz), 7.28 (d, 2H, J = 8.30 Hz). 13
C
NMR (150 MHz, CDCl3): ı 11.24, 12.28, 20.76, 21.15, 22.67, 23.35,
28.63, 29.93, 30.88, 31.63, 34.93, 35.84, 37.38, 42.97, 44.51, 45.58,
47.38, 48.50, 53.08, 54.44, 63.75, 84.77, 122.57, 128.11, 129.02,
135.16, 135.90, 145.12. Anal. Calcd. for C32H48N2O: C 80.62, H 10.15,
N 5.88; Found C 80.39, H 10.01, N 5.99.
2.5.10. 16-(4-nitro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-ol (9c)
Yellow solid. Yield 45%. m.p. 221–223 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.72 (s, 3H, CH3-19), 0.85 (s, 3H, CH3-18), 2.59 (br, 4H,
NCH2), 4.10 (s, 1H, C17-␣H), 6.62 (d, 1H, CH, J = 2.28 Hz), 7.52 (d,
2H, J = 8.88 Hz), 8.21 (d, 2H, J = 8.88 Hz). 13C NMR (150 MHz, CDCl3):
ı 11.22, 12.38, 20.66, 23.18, 27.93, 28.69, 31.23, 31.83, 34.67, 34.99,
35.95, 36.36, 37.42, 43.17, 45.32, 48.51, 51.84, 54.67, 64.35, 85.08,
121.40, 123.75, 128.53, 144.51, 145.65, 152.15. Anal. Calcd. for
2.5.5. 16-(4-methoxy-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-ol (8b)
White solid. Yield 41%. m.p. 204–206 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.53 (s, 4H,
NCH2), 3.83 (s, 3H, OCH3), 4.06 (d, 1H, C17-␣H, J = 8.52 Hz), 6.46 (s,
1H, CH), 6.90 (d, 2H, J = 8.80 Hz), 7.35 (d, 2H, J = 8.76 Hz). 13C NMR
(150 MHz, CDCl3): ı 11.17, 12.46, 20.80, 24.12, 24.88, 26.42, 28.88,
30.68, 30.90, 31.85, 35.06, 36.13, 36.43, 38.02, 43.03, 46.13, 48.58,
50.36, 54.70, 55.26, 64.54, 85.02, 113.77, 122.16, 129.39, 130.80,
143.93, 158.01. Anal. Calcd. for C32H47NO2: C 80.45, H 9.92, N 2.93;
Found C 80.27, H 9.88, N 3.02.
C30H42N2O3: C 75.28, H 8.84, N 5.85; Found C 75.31, H 8.77, N 5.93.
2.5.11. 16-(4-nitro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-ol (10c)
Yellow solid. Yield 41%. m.p. 212–214 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.71 (s, 3H, CH3-19), 0.87 (s, 3H, CH3-18), 2.42 (s, 4H,
NCH2), 3.72–3.74 (m, 4H, OCH2), 4.09 (s, 1H, C17-␣H), 6.62 (d, 1H,
CH, J = 2.28 Hz), 7.52 (d, 2H, J = 8.88 Hz), 8.21 (d, 2H, J = 8.88 Hz). 13
C
NMR (150 MHz, CDCl3): ı 11.23, 12.24, 20.39, 23.74, 28.41, 30.62,
31.23, 31.73, 32.84, 34.97, 36.28, 36.39, 39.42, 43.19, 48.48, 50.84,
54.55, 59.35, 67.41, 85.16, 121.39, 123.75, 128.54, 144.50, 145.65,
152.10. Anal. Calcd. for C30H42N2O4: C 72.84, H 8.56, N 5.66; Found
C 72.66, H 8.41, N 5.39.
2.5.6. 16-(4-methoxy-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-ol (9b)
White solid. Yield 41%. m.p. 178–180 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.67 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.30 (s, 3H,
N-CH3), 2.57 (br, 4H, NCH2), 3.83 (s, 3H, OCH3), 4.03 (d, 1H, C17
-
␣H, J = 8.52 Hz), 6.44 (s, 1H, CH), 6.88 (d, 2H, J = 8.72 Hz), 7.33 (d,
2H, J = 8.72 Hz). 13C NMR (150 MHz, CDCl3): ı 11.17, 12.37, 20.73,
23.18, 27.67, 28.75, 30.88, 31.82, 34.42, 35.03, 35.92, 36.43, 37.38,
43.01, 45.32, 48.60, 51.73, 54.67, 55.25, 64.36, 84.98, 113.76, 122.44,
129.39, 130.82, 143.93, 157.99. Anal. Calcd. for C31H45NO2: C 80.30,
H 9.78, N 3.02; Found C 80.22, H 9.91, N 2.89.
2.5.12. 16-(4-nitro-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-ol (11c)
Yellow solid. Yield 40%. m.p. 244–246 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.71 (s, 3H, CH3-19), 0.87 (s, 3H, CH3-18), 2.30 (s, 3H,
N-CH3), 2.48–2.65 (m, 8H, NCH2), 4.09 (s, 1H, C17-␣H), 6.62 (d, 1H,
CH, J = 2.00 Hz), 7.52 (d, 2H, J = 8.80 Hz), 8.21 (d, 2H, J = 8.80 Hz). 13
C
NMR (150 MHz, CDCl3): ı 11.24, 12.42, 20.70, 24.51, 28.74, 31.22,
31.79, 34.97, 36.03, 37.78, 43.17, 45.87, 46.01, 48.47, 49.24, 54.63,
55.44, 63.79, 85.06, 121.43, 123.75, 128.54, 144.48, 145.66, 152.06.
Anal. Calcd. for C31H45N3O3: C 73.34, H 8.93, N 8.28; Found C 73.17,
H 8.76, N 8.51.
2.5.7. 16-(4-methoxy-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-ol (10b)
White solid. Yield 47%. m.p. 173–175 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.86 (s, 3H, CH3-18), 2.59 (s, 4H,
NCH2), 3.73–3.75 (m, 4H, OCH2), 3.83 (s, 3H, OCH3), 4.06 (d, 1H,
C17-␣H, J = 8.88 Hz), 6.47 (s, 1H, CH), 6.90 (d, 2H, J = 8.72 Hz), 7.35
(d, 2H, J = 8.72 Hz). 13C NMR (150 MHz, CDCl3): ı 11.18, 12.41, 20.78,
24.50, 28.85, 30.89, 31.12, 31.81, 35.03, 36.04, 37.70, 43.02, 45.76,
46.14, 48.55, 50.05, 54.65, 55.26, 64.12, 67.35, 85.00, 113.76, 122.18,
2.5.13. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-ol (8d)
White solid. Yield 49%. m.p. 217–219 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.68 (s, 3H, CH3-19), 0.80 (s, 3H, CH3-18), 2.51 (s, 4H,

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NCH2), 3.86 (s, 9H, OCH3), 4.09 (d, 1H, C17-␣H, J = 9.00 Hz), 6.44
(d, 1H, CH, J = 2.00 Hz), 6.62 (s, 2H). 13C NMR (150 MHz, CDCl3): ı
11.20, 12.44, 20.77, 24.08, 24.86, 26.38, 28.84, 30.64, 30.78, 31.82,
35.00, 36.12, 36.40, 38.00, 43.01, 46.10, 48.53, 50.34, 54.68, 56.05,
60.91, 64.51, 84.93, 105.48, 122.83, 133.72, 136.70, 145.63, 152.99.
Anal. Calcd. for C34H51NO4: C 75.94, H 9.56, N 2.60; Found C 75.88,
H 9.41, N 2.39.
131.83, 136.39, 147.15. Anal. Calcd. for C30H42ClNO: C 76.97, H 9.04,
N 2.99; Found C 77.11, H 8.89, N 2.74.
2.5.19. 16-(4-chloro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-ol (10e)
White solid. Yield 41%. m.p. 224–226 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.70 (s, 3H, CH3-19), 0.84 (s, 3H, CH3-18), 2.58 (s, 4H,
NCH2), 3.82–3.84 (m, 4H, OCH2), 4.06 (d, 1H, C17-␣H, J = 8.64 Hz),
6.48 (d, 1H, CH, J = 2.24 Hz), 7.30 (s, 4H). 13C NMR (150 MHz,
CDCl3): ı 11.20, 12.43, 20.66, 28.13, 28.77, 30.88, 31.79, 34.75,
35.13, 35.97, 36.29, 37.51, 43.12, 45.29, 48.71, 51.78, 54.66, 64.21,
85.12, 121.71, 128.55, 129.24, 131.79, 136.28, 147.22. Anal. Calcd.
for C30H42ClNO2: C 74.43, H 8.74, N 2.89; Found C 74.51, H 8.82, N
2.97.
2.5.14. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-ol (9d)
White solid. Yield 48%. m.p. 186–188 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.83 (s, 3H, CH3-18), 2.56 (br, 4H,
NCH2), 3.87 (s, 9H, OCH3), 4.04 (d, 1H, C17-␣H, J = 8.28 Hz), 6.43 (d,
1H, CH, J = 2.00 Hz), 6.62 (s, 2H). 13C NMR (150 MHz, CDCl3): ı 9.77,
10.96, 19.29, 21.77, 26.58, 27.35, 29.38, 30.44, 33.35, 33.60, 34.54,
34.99, 36.02, 41.61, 43.91, 47.15, 50.43, 53.29, 54.68, 59.52, 62.93,
83.64, 104.09, 121.47, 132.27, 135.36, 144.23, 151.61. Anal. Calcd.
for C33H49NO4: C 75.68, H 9.43, N 2.67; Found C 75.51, H 9.19, N
2.59.
2.5.20. 16-(4-chloro-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-ol (11e)
White solid. Yield 41%. m.p. 229–231 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.31 (s, 3H,
N-CH3), 2.51–2.63 (m, 8H, NCH2), 4.05 (s, 1H, C17-␣H), 6.48 (d,
1H, CH, J = 2.26 Hz), 7.30 (s, 4H). 13C NMR (150 MHz, CDCl3): ı
11.19, 12.42, 20.74, 24.47, 28.78, 30.90, 31.18, 31.79, 35.00, 36.02,
37.77, 43.05, 45.87, 46.00, 48.50, 49.22, 54.64, 55.35, 63.88, 84.94,
121.76, 128.46, 129.39, 131.84, 136.38, 147.08. Anal. Calcd. for
2.5.15. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-ol (10d)
White solid. Yield 48%. m.p. 241–243 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.56 (s, 4H,
C31H45ClN2O: C 74.89, H 9.12, N 5.63; Found C 74.69, H 9.03, N
NCH2), 3.72–3.75 (m, 4H, OCH2), 3.84 (s, 9H, OCH3), 4.03 (d, 1H, C17
-
5.51.
␣H, J = 8.28 Hz), 6.43 (d, 1H, CH, J = 1.60 Hz), 6.62 (s, 2H). 13C NMR
(150 MHz, CDCl3): ı 11.19, 12.38, 20.73, 24.44, 28.79, 30.75, 31.08,
31.76, 34.96, 36.01, 36.36, 37.66, 42.98, 45.72, 48.49, 50.01, 54.61,
56.04, 60.89, 64.07, 67.30, 84.91, 105.48, 122.85, 133.68, 136.70,
145.50, 152.97. Anal. Calcd. for C33H49NO5: C 73.43, H 9.15, N 2.60;
Found C 73.21, H 9.01, N 2.81.
2.5.21. 16-(4-bromo-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-ol (8f)
White solid. Yield 44%. m.p. 210–212 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.54 (s, 4H,
NCH2), 4.05 (d, 1H, C17-␣H, J = 8.80 Hz), 6.46 (s, 1H, CH), 7.26 (d,
2H, J = 8.44 Hz), 7.46 (d, 2H, J = 8.44 Hz). 13C NMR (150 MHz, CDCl3):
ı 11.20, 12.45, 20.75, 24.07, 24.81, 26.31, 28.81, 30.57, 30.92, 31.81,
35.00, 36.11, 37.98, 43.04, 46.09, 48.50, 50.34, 54.65, 64.57, 84.93,
119.96, 121.77, 129.72, 131.39, 136.83, 147.34. Anal. Calcd. for
2.5.16. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-(4-
methylpiperazinyl)-5˛-
androstan-17ˇ-ol (11d)
White solid. Yield 48%. m.p. 210–212 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.68 (s, 3H, CH3-19), 0.80 (s, 3H, CH3-18), 2.28 (s, 3H,
N-CH3), 2.47–2.62 (m, 8H, NCH2), 3.86 (s, 9H, OCH3), 4.04 (d, 1H,
C17-␣H, J = 8.28 Hz), 6.44 (d, 1H, CH, J = 1.60 Hz), 6.62 (s, 2H).
13C NMR (150 MHz, CDCl3): ı 10.31, 11.55, 19.88, 23.67, 27.94,
29.92, 30.50, 30.94, 34.14, 35.18, 35.51, 36.94, 42.15, 45.02, 45.19,
47.65, 48.46, 53.80, 54.67, 55.21, 60.06, 62.95, 84.16, 104.63, 122.02,
132.79, 135.91, 144.72, 152.15. Anal. Calcd. for C34H52N2O4: C
73.87, H 9.48, N 5.07; Found C 73.77, H 9.62, N 4.91.
C31H44BrNO: C 70.71, H 8.42, N 2.66; Found C 70.59, H 8.31, N 2.88.
2.5.22. 16-(4-bromo-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-ol (9f)
White solid. Yield 46%. m.p. 203–205 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.84 (s, 3H, CH3-18), 2.58 (br,
4H, NCH2), 4.04 (s, 1H, C17-␣H), 6.46 (s, 1H, CH), 7.26 (d, 2H,
J = 8.60 Hz), 7.46 (d, 2H, J = 8.62 Hz). 13C NMR (150 MHz, CDCl3): ı
11.18, 12.37, 20.69, 23.19, 27.83, 28.73, 30.92, 31.82, 35.00, 35.93,
37.40, 43.04, 45.31, 46.17, 48.53, 51.80, 54.67, 64.36, 85.00, 119.98,
121.78, 129.72, 131.39, 136.81, 147.35. Anal. Calcd. forC30H42BrNO:
C 70.30, H 8.26, N 2.73; Found C 70.08, H 8.41, N 2.59.
2.5.17. 16-(4-chloro-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-ol (8e)
White solid. Yield 48%. m.p. 225–227 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.53 (s, 4H,
NCH2), 4.06 (d, 1H, C17-␣H, J = 8.88 Hz), 6.48 (d, 1H, CH, J = 2.28 Hz),
7.30 (s, 4H). 13C NMR (150 MHz, CDCl3): ı 11.88, 12.45, 20.76, 24.17,
24.90, 26.46, 28.84, 30.68, 30.91, 31.84, 35.03, 36.13, 38.03, 43.06,
46.13, 48.52, 50.38, 54.69, 64.55, 84.98, 121.75, 128.46, 129.39,
131.84, 136.39, 147.12. Anal. Calcd. for C31H44ClNO: C 77.22, H 9.20,
N 2.91; Found C 77.03, H 9.01, N 3.03.
2.5.23. 16-(4-bromo-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-ol (10f)
White solid. Yield 46%. m.p. 241–243 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.68 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.57 (s, 4H,
NCH2), 3.82–3.85 (m, 4H, OCH2), 4.04 (s, 1H, C17-␣H), 6.46 (s, 1H,
CH), 7.26 (d, 2H, J = 8.60 Hz), 7.46 (d, 2H, J = 8.62 Hz). 13C NMR
(150 MHz, CDCl3): ı 11.19, 12.51, 20.77, 20.11, 24.81, 26.09, 28.91,
30.58, 30.99, 31.77, 35.02, 36.15, 37.99, 43.01, 46.10, 48.48, 50.31,
54.62, 64.55, 84.90, 119.98, 121.73, 129.68, 131.41, 136.79, 147.37.
Anal. Calcd. for C30H42BrNO2: C 68.17, H 8.01, N 2.65; Found C 68.03,
H 7.96, N 2.78.
2.5.18. 16-(4-chloro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-ol (9e)
White solid. Yield 41%. m.p. 224–226 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.70 (s, 3H, CH3-19), 0.84 (s, 3H, CH3-18), 2.58 (br, 4H,
NCH2), 4.06 (d, 1H, C17-␣H, J = 8.64 Hz), 6.48 (d, 1H, CH, J = 2.24 Hz),
7.30 (s, 4H). 13C NMR (150 MHz, CDCl3): ı 11.18, 12.38, 20.70, 23.19,
27.99, 28.75, 30.92, 31.84, 34.75, 35.02, 35.95, 36.40, 37.44, 43.06,
45.33, 48.55, 51.86, 54.70, 64.39, 85.00, 121.74, 128.46, 129.39,
2.5.24. 16-(4-bromo-benzylidene)-3ˇ-(4-methylpiperazinyl)-
5˛-androstan-17ˇ-ol (11f)
White solid. Yield 46%. m.p. 235–237 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.60 (s, 3H, CH3-19), 0.73 (s, 3H, CH3-18), 2.22 (s, 3H, N-
CH3), 2.43–2.54 (m, 8H, NCH2), 3.94 (s, 1H, C17-␣H), 6.37 (s, 1H,

H. Guo et al. / Steroids 76 (2011) 709–723
717
CH), 7.17 (d, 2H, J = 8.48 Hz), 7.37 (d, 2H, J = 8.44 Hz). 13C NMR
(150 MHz, CDCl3): ı 11.22, 12.42, 20.74, 24.47, 28.78, 30.90, 31.18,
31.79, 34.99, 36.02, 37.77, 43.04, 45.85, 46.00, 48.50, 49.20, 54.63,
55.35, 63.85, 84.90, 119.97, 121.79, 129.73, 131.40, 136.84, 147.34.
Anal. Calcd. for C31H45BrN2O: C 68.75, H 8.37, N 5.17; Found C 68.79,
H 8.51, N 5.23.
2.5.30. 16-(furan-2-ylmethylene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-ol (9h)
White solid. Yield 44%. m.p. 220–222 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.67 (s, 3H, CH3-19), 0.84 (s, 3H, CH3-18), 2.58 (br, 4H,
NCH2), 4.07 (d, 1H, C17-␣H, J = 8.60 Hz), 6.22 (d, 1H, CH, J = 3.24 Hz),
6.42 (s, 2H), 7.39 (s, 1H). 13C NMR (150 MHz, CDCl3): ı 11.15, 12.38,
20.72, 23.17, 27.90, 28.78, 30.97, 31.83, 34.68, 35.02, 35.93, 36.41,
37.44, 43.38, 45.34, 48.37, 51.80, 54.71, 64.36, 84.52, 107.43, 111.21,
111.62, 141.14, 145.42, 153.82. Anal. Calcd. for C28H41NO2: C 79.39,
H 9.76, N 3.31; Found C 79.55, H 9.59, N 3.43.
2.5.25. 16-(4-fluoro-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-ol (8g)
White solid. Yield 46%. m.p. 188–190 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.69 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.57 (s, 4H,
NCH2), 4.04 (s, 1H, C17-␣H), 6.48 (d, 1H, CH, J = 2.08 Hz), 7.02
(t, 2H), 7.35 (q, 2h). 13C NMR (150 MHz, CDCl3): ı 11.24, 12.41,
20.77, 23.82, 24.65, 26.00, 28.80, 30.31, 31.78, 34.99, 36.07, 37.91,
43.01, 45.97, 46.03, 48.55, 50.14, 54.65, 64.46, 84.63, 115.01, 115.22,
121.62, 129.59, 134.24, 145.87, 159.92, 162.37. Anal. Calcd. for
2.5.31. 16-(furan-2-ylmethylene)-3ˇ-morpholino-5˛-
androstan-17ˇ-ol (10h)
White solid. Yield 49%. m.p. 220–222 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.68 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.58 (s, 4H,
NCH2), 3.73–3.76 (m, 4H, OCH2), 4.06 (s, 1H, C17-␣H), 6.23 (d, 1H,
CH, J = 3.20 Hz), 6.42 (s, 2H), 7.39 (s, 1H). 13C NMR (150 MHz,
CDCl3): ı 11.15, 12.41, 20.77, 24.48, 28.84, 30.96, 31.11, 31.78,
35.01, 36.03, 37.70, 43.38, 45.78, 48.31, 50.04, 54.66, 64.11, 67.33,
84.53, 107.47, 111.22, 111.67, 141.18, 145.27, 153.80. Anal. Calcd.
for C28H41NO3: C 76.50, H 9.40, N 3.19; Found C 76.22, H 9.51, N
3.29.
C31H44FNO: C 79.95, H 9.52, N 3.01; Found C 79.80, H 9.38, N
3.22.
2.5.26. 16-(4-fluoro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-ol (9g)
White solid. Yield 46%. m.p. 240–242 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.60 (s, 3H, CH3-19), 0.76 (s, 3H, CH3-18), 2.37–2.39 (m,
4H, NCH2), 3.96 (s, 1H, C17-␣H, J = 8.64 Hz), 6.39 (d, 1H, CH), 6.94 (t,
2H), 7.27 (q, 2 h). 13C NMR (150 MHz, CDCl3): ı 11.26, 12.45, 20.87,
23.90, 26.11, 28.91, 30.13, 31.82, 34.96, 36.10, 37.92, 43.05, 45.88,
46.12, 48.50,50.12, 54.66, 64.45, 84.60, 115.04, 115.21, 121.56,
129.60, 134.31, 145.93, 159.95, 162.40. Anal. Calcd. for C30H42FNO:
C 79.78, H 9.37, N 3.10; Found C 79.61, H 9.45, N 3.01.
2.5.32. 16-(furan-2-ylmethylene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-ol (11h)
White solid. Yield 40%. m.p. 239–241 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.67 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.30 (s, 3H,
N-CH3), 2.48–2.62 (m, 8H, NCH2), 4.06 (s, 1H, C17-␣H), 6.22 (d,
1H, CH, J = 3.20 Hz), 6.42 (s, 2H), 7.39 (s, 1H). 13C NMR (150 MHz,
CDCl3): ı 11.14, 12.43, 14.20, 20.77, 21.06, 24.55, 28.84, 30.97,
31.32, 31.81, 35.03, 36.04, 37.83, 43.39, 45.93, 46.06, 48.33, 49.30,
54.70, 55.53, 60.40, 63.84, 84.57, 107.49, 111.23, 111.67, 141.20,
145.36, 153.83. Anal. Calcd. for C29H44N2O2: C 76.95, H 9.80, N 6.19;
Found C 76.80, H 9.91, N 6.11.
2.5.27. 16-(4-fluoro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-ol (10g)
White solid. Yield 46%. m.p. 183–185 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.61 (s, 3H, CH3-19), 0.73 (s, 3H, CH3-18), 2.46–2.49 (m,
4H, NCH2), 3.64–3.66 (m, 4H, OCH2), 3.96 (s, 1H, C17-␣H), 6.40 (d,
1H, CH), 6.94 (t, 2H), 7.26 (q, 2h). 13C NMR (150 MHz, CDCl3): ı
11.31, 12.52, 20.79, 23.88, 26.13, 29.09, 30.15, 31.88, 34.95, 36.01,
37.99, 43.03, 45.99, 36.15, 48.01, 50.11, 54.84, 64.44, 84.55, 115.01,
115.22, 121.55, 129.49, 134.13, 145.90, 159.99, 162.38. Anal. Calcd.
for C30H42FNO2: C 77.05, H 9.05, N 3.00; Found C 76.88, H 8.91, N
3.21.
2.6. General procedure for the synthesis of compounds 12a–h,
13a–h, 14a–h, 15a–h
To a solution of compounds 8a–h, 9a–h, 10a–h, 11a–h (5 mmol)
in CH2Cl2 was added acetic anhydride (10 mmol, 2 equiv) and 4-
dimethylaminopyridine (5 mmol, 1 equiv). The mixture was stirred
at 40 ◦C until the reaction was completed (monitored by TLC). The
reaction was concentrated in reduced pressure. The crude product
was dissolved in CH2Cl2, the organic layer was washed with brine
and dried over anhydrous Na2SO4, filtered. The filtrate was concen-
trated in reduced pressure to afford a crude product. The products
were purified by flash column chromatography to yield compound
12a–h, 13a–h, 14a–h, 15a–h.
2.5.28. 16-(4-fluoro-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-ol (11g)
White solid. Yield 46%. m.p. 213–215 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.61 (s, 3H, CH3-19), 0.73 (s, 3H, CH3-18), 2.21 (s, 3H,
N-CH3), 2.40–2.55 (m, 8H, NCH2), 3.96 (s, 1H, C17-␣H), 6.40 (d, 1H,
CH), 6.94 (t, 2H), 7.26 (q, 2H). 13C NMR (150 MHz, CDCl3): ı 11.20,
12.42, 14.19, 20.75, 24.54, 28.80, 30.77, 31.24, 31.81, 35.01, 36.03,
37.81, 43.03, 45.88, 46.03, 48.54, 49.23, 54.67, 55.47, 60.40, 63.82,
84.81, 115.27, 121.70, 129.68, 134.13, 145.83, 159.99, 162.43. Anal.
Calcd. for C31H45FN2O: C 77.46, H 9.44, N 5.83; Found C 77.63, H
9.19, N 5.77.
2.6.1. 16-(4-methyl-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-yl acetate (12a)
White solid. Yield 95%. m.p. 171–173 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.80 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.35 (s, 3H, Ph-CH3), 2.52 (s, 4H, NCH2), 5.37 (s, 1H, C17
-
␣H), 6.18 (d, 1H, CH, J = 2.12 Hz), 7.15 (d, 2H, J = 8.00 Hz), 7.20 (d,
2H, J = 8.08 Hz). 13C NMR (150 MHz, CDCl3): ı 12.30, 12.42, 20.66,
21.20, 24.20, 24.94, 26.52, 28.86, 30.75, 31.00, 31.74, 34.89, 36.07,
37.97, 42.97, 46.03, 48.88, 50.40, 54.40, 64.53, 84.63, 123.36, 128.21,
129.00, 134.85, 136.08, 139.94, 171.10. Anal. Calcd. for C34H49NO2:
C 81.06, H 9.80, N 2.78; Found C 81.22, H 9.71, N 2.90. MS (m/z):
504.7[M+1]+.
2.5.29. 16-(furan-2-ylmethylene)-3ˇ-piperidino-5˛-
androstan-17ˇ-ol (8h)
White solid. Yield 41%. m.p. 267–269 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.67 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.54 (s, 4H,
NCH2), 4.06 (s, 1H, C17-␣H), 6.23 (d, 1H, CH, J = 3.24 Hz), 6.42
(s, 2H), 7.39 (s, 1H). 13C NMR (150 MHz, CDCl3): ı 11.15, 12.43,
20.79, 24.51, 28.89, 30.97, 31.12, 31.77, 35.10, 36.08, 36.33, 37.69,
43.39, 45.77, 48.35, 50.00, 54.69, 64.13, 67.33, 84.55, 107.49, 111.29,
111.66, 141.14, 145.22, 153.80. Anal. Calcd. for C29H43NO2: C 79.59,
H 9.90, N 3.20; Found C 79.67, H 9.79, N 3.08.
2.6.2. 16-(4-methyl-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-yl acetate (13a)
White solid. Yield 92%. m.p. 188–190 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.83 (s, 3H, CH3-18), 2.21 (s, 3H,

718
H. Guo et al. / Steroids 76 (2011) 709–723
OCOCH3), 2.35 (s, 3H, Ph-CH3), 2.57 (br, 4H, NCH2), 5.37 (s, 1H, C17
-
OCOCH3), 2.59 (s, 4H, NCH2), 3.74–3.77 (m, 4H, OCH2), 3.83 (s, 3H,
OCH3), 5.36 (s, 1H, C17-␣H), 6.16 (d, 1H, CH, J = 2.16 Hz), 6.89 (d,
2H, J = 8.80 Hz), 7.31 (d, 2H, J = 8.80 Hz). 13C NMR (150 MHz, CDCl3):
ı 12.31, 12.38, 20.65, 21.25, 24.48, 28.80, 30.92, 31.09, 31.72, 34.88,
36.00, 37.64, 42.98, 45.67, 48.91, 50.05, 54.36, 55.25, 64.11, 67.33,
84.67, 113.71, 122.93, 129.48, 130.48, 138.56, 158.11, 171.27. Anal.
Calcd. for C33H47NO4: C 75.97, H 9.08, N 2.68; Found C 75.88, H
8.97, N 2.56. MS (m/z): 522.6[M+1]+.
␣H), 6.18 (d, 1H, CH, J = 2.00 Hz), 7.15 (d, 2H, J = 8.00 Hz), 7.26 (d,
2H, J = 8.04 Hz). 13C NMR (150 MHz, CDCl3): ı 12.28, 12.33, 20.58,
21.17, 21.21, 23.18, 27.93, 28.74, 30.98, 31.74, 34.70, 34.89, 35.89,
36.55, 37.37, 42.97, 45.23, 48.92, 51.83, 54.42, 64.35, 84.65, 123.35,
128.19, 128.99, 134.83, 136.13, 139.96, 171.19. Anal. Calcd. for
C33H47NO2: C 80.93, H 9.67, N 2.86; Found C 80.77, H 9.58, N 2.99.
MS (m/z): 490.6[M+1]+.
2.6.3. 16-(4-methyl-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-yl acetate (14a)
2.6.8. 16-(4-methoxy-benzylidene)-3ˇ-(4-methylpiperazinyl)-
5˛-androstan-17ˇ-yl acetate (15b)
White solid. Yield 96%. m.p. 96–98 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.83 (s, 3H, CH3-18), 2.18 (s, 3H,
OCOCH3), 2.27 (s, 3H, NCH3), 2.32 (s, 3H, Ph-CH3), 2.57 (s, 4H,
NCH2), 3.73–3.75 (m, 4H, OCH2) 5.37 (s, 1H, C17-␣H), 6.18 (d, 1H,
CH, J = 2.02 Hz), 7.15 (d, 2H, J = 7.88 Hz), 7.26 (d, 2H, J = 8.00 Hz).
13C NMR (150 MHz, CDCl3): ı 11.55, 12.29, 12.37, 20.64, 21.17,
21.21, 24.50, 28.81, 30.97, 31.12, 31.71, 34.88, 36.00, 37.65, 42.97,
45.68, 46.21, 48.88, 50.06, 54.37, 64.08, 67.37, 84.62, 123.38, 128.19,
129.00, 134.82, 136.15, 139.91, 171.19. Anal. Calcd. for C33H47NO3:
C 78.37, H 9.37, N 2.77; Found C 78.52, H 9.45, N 2.59. MS (m/z):
506.6[M+1]+.
White solid. Yield 91%. m.p. 186–188 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.30 (s, 3H, Ph-CH3), 2.50–2.67 (m, 8H, NCH2), 3.82 (s, 3H,
OCH3), 5.36 (s, 1H, C17-␣H), 6.15 (d, 1H, CH, J = 2.16 Hz), 6.89 (d,
2H, J = 8.80 Hz), 7.31 (d, 2H, J = 8.76 Hz). 13C NMR (150 MHz, CDCl3):
ı 12.30, 12.39, 20.63, 21.23, 24.53, 28.79, 30.91, 31.27, 31.72, 34.88,
35.98, 36.52, 37.75, 42.96, 45.79, 46.05, 48.90, 49.30, 54.37, 55.22,
55.52, 63.80, 84.67, 113.69, 122.89, 129.47, 130.48, 138.58, 158.09,
171.21. Anal. Calcd. for C34H50N2O3: C 76.36, H 9.42, N 5.24; Found
C 76.47, H 9.60, N 5.11. MS (m/z): 535.7[M+1]+.
2.6.9. 16-(4-nitro-benzylidene)-3ˇ-piperidino-5˛-
2.6.4. 16-(4-methyl-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-yl acetate (15a)
androstan-17ˇ-yl acetate (12c)
Yellow solid. Yield 90%. m.p. 190–192 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.78 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.23 (s, 3H,
OCOCH3), 2.52 (s, 4H, NCH2), 5.39 (s, 1H, C17-␣H), 6.28 (d, 1H, CH,
J = 2.20 Hz), 7.49 (d, 2H, J = 8.84 Hz), 8.21 (d, 2H, J = 8.84 Hz). 13C NMR
(150 MHz, CDCl3): ı 12.27, 12.38, 20.56, 21.11, 24.11, 24.86, 26.42,
28.72, 30.63, 31.28, 31.72, 34.84, 36.06, 37.92, 43.09, 45.99, 48.75,
50.35, 54.36, 64.44, 84.52, 121.88, 123.69, 128.65, 144.09, 145.81,
146.98, 170.97. Anal. Calcd. for C33H46N2O4: C 74.12, H 8.67, N 5.24;
Found C 74.29, H 8.88, N 5.09. MS (m/z): 535.6[M+1]+.
White solid. Yield 95%. m.p. 180–182 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.75 (s, 3H, CH3-19), 0.79 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.35 (s, 3H, Ph-CH3), 2.46–2.60 (m, 8H, NCH2), 5.34 (s, 1H,
C17-␣H), 6.16 (d, 1H, CH, J = 2.06 Hz), 7.13 (d, 2H, J = 7.96 Hz), 7.24
(d, 2H, J = 8.12 Hz). 13C NMR (150 MHz, CDCl3): ı 12.27, 12.37, 20.62,
21.17, 24.52, 28.79, 30.94, 31.27, 31.68, 34.85, 35.95, 37.73, 42.93,
45.76, 46.03, 48.83, 49.27, 54.34, 55.51, 63.75, 84.58, 123.34, 128.18,
128.97, 134.81, 136.05, 139.88, 171.05. Anal. Calcd. for C34H50N2O2:
C 78.72, H 9.71, N 5.40; Found C 78.88, H 9.60, N 5.19. MS (m/z):
519.6[M+1]+.
2.6.10. 16-(4-nitro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-yl acetate (13c)
2.6.5. 16-(4-methoxy-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-yl acetate (12b)
Yellow solid. Yield 90%. m.p. 143–145 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.78 (s, 3H, CH3-19), 0.83 (s, 3H, CH3-18), 2.22 (s, 3H,
OCOCH3), 2.57 (br, 4H, NCH2), 5.38 (s, 1H, C17-␣H), 6.28 (d, 1H, CH,
J = 2.20 Hz), 7.49 (d, 2H, J = 8.80 Hz), 8.20 (d, 2H, J = 8.76 Hz). 13C NMR
(150 MHz, CDCl3): ı 12.25, 12.30, 20.49, 21.10, 23.15, 27.88, 28.62,
31.27, 31.71, 34.64, 34.82, 35.86, 37.34, 43.08, 45.18, 48.76, 51.77,
54.36, 64.23, 84.52, 121.89, 123.69, 128.66, 144.09, 145.80, 146.98,
171.00. Anal. Calcd. for C32H44N2O4: C 73.81, H 8.52, N 5.38; Found
C 73.99, H 8.49, N 5.17. MS (m/z): 521.6[M+1]+.
White solid. Yield 95%. m.p. 190–192 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.80 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.53 (s, 4H, NCH2), 3.81 (s, 3H, OCH3), 5.36 (s, 1H, C17-␣H),
6.15 (d, 1H, CH, J = 2.08 Hz), 6.89 (d, 2H, J = 8.44 Hz), 7.31 (d, 2H,
J = 8.76 Hz). 13C NMR (150 MHz, CDCl3): ı 12.30, 12.41, 20.65, 21.24,
24.05, 24.79, 26.29, 28.81, 30.61, 30.93, 31.74, 34.89, 36.07, 37.91,
42.98, 46.00, 48.92, 50.34, 54.39, 55.24, 64.55, 84.68, 113.69, 122.88,
129.45, 130.48, 138.60, 158.08, 171.23. Anal. Calcd. for C34H49NO3:
C 78.57, H 9.50, N 2.69; Found C 78.44, H 9.37, N 2.81. MS (m/z):
520.6[M+1]+.
2.6.11. 16-(4-nitro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-yl acetate (14c)
Yellow solid. Yield 95%. m.p. 122–124 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.78 (s, 3H, CH3-19), 0.86 (s, 3H, CH3-18), 2.23 (s, 3H,
OCOCH3), 2.42 (s, 4H, NCH2), 3.72–3.74 (m, 4H, OCH2), 5.39 (s,
1H, C17-␣H), 6.29 (d, 1H, CH, J = 2.16 Hz), 7.50 (d, 2H, J = 8.80 Hz),
8.21 (d, 2H, J = 8.76 Hz). 13C NMR (150 MHz, CDCl3): ı 12.19, 12.29,
20.25, 21.13, 23.70, 28.37, 30.69, 31.28, 31.63, 32.80, 34.84, 36.36,
39.34, 43.12, 48.73, 50.86, 54.22, 59.36, 67.38, 84.59, 121.87, 123.71,
128.67, 144.12, 145.81, 147.06, 171.03. Anal. Calcd. for C32H44N2O5:
C 71.61, H 8.26, N 5.22; Found C 71.48, H 8.44, N 5.06. MS (m/z):
537.6[M+1]+.
2.6.6. 16-(4-methoxy-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-yl acetate (13b)
White solid. Yield 92%. m.p. 205–207 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.19 (s, 3H,
OCOCH3), 2.62 (br, 4H, NCH2), 3.81 (s, 3H, OCH3), 5.34 (s, 1H, C17
-
␣H), 6.13 (d, 1H, CH, J = 2.08 Hz), 6.87 (d, 2H, J = 8.56 Hz), 7.30 (d,
2H, J = 8.76 Hz). 13C NMR (150 MHz, CDCl3): ı 12.29, 12.42, 20.59,
21.23, 23.18, 26.88, 28.58, 30.90, 31.65, 33.57, 34.82, 36.51, 37.17,
42.95, 45.20, 48.92, 51.56, 54.27, 55.23, 64.41, 84.65, 113.70, 113.75,
122.93, 129.68, 130.45, 138.50, 158.11, 171.26. Anal. Calcd. for
C
33H47NO3: C 78.37, H 9.37, N 2.77; Found C 78.61, H 9.53, N 2.59.
2.6.12. 16-(4-nitro-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-yl acetate (15c)
MS (m/z): 506.6[M+1]+.
Yellow solid. Yield 98%. m.p. 177–179 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.76 (s, 3H, CH3-19), 0.79 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.28 (s, 3H, Ph-CH3), 2.48–2.61 (m, 8H, NCH2), 5.37 (s, 1H,
2.6.7. 16-(4-methoxy-benzylidene)-3ˇ-morpholino-5˛
androstan-17ˇ-yl acetate (14b)
White solid. Yield 88%. m.p. 217–219 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.21 (s, 3H,
C17-␣H), 6.27 (d, 1H, CH, J = 2.08 Hz), 7.47 (d, 2H, J = 8.84 Hz), 8.18
(d, 2H, J = 8.84 Hz). 13C NMR (150 MHz, CDCl3): ı 12.22, 12.33, 20.52,

H. Guo et al. / Steroids 76 (2011) 709–723
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21.05, 24.45, 28.66, 31.17, 31.22, 31.65, 34.78, 35.93, 37.69, 43.04,
45.73, 45.95, 48.70, 49.19, 54.31, 55.41, 63.69, 84.44, 121.86, 123.64,
128.63, 144.03, 145.78, 146.92, 170.87. Anal. Calcd. for C33H47N3O4:
C 72.10, H 8.62, N 7.64; Found C 72.31, H 8.47, N 7.49. MS (m/z):
550.6[M+1]+.
84.56, 122.37, 128.43, 129.46, 132.03, 136.08, 141.96, 171.13. Anal.
Calcd. for C33H46ClNO2: C 75.61, H 8.85, N 2.67; Found C 75.44, H
8.67, N 2.59. MS (m/z): 524.5[M+1]+.
2.6.18. 16-(4-chloro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-yl acetate (13e)
2.6.13. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-yl acetate (12d)
White solid. Yield 95%. m.p. 185–187 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.68 (s, 3H, CH3-19), 0.74 (s, 3H, CH3-18), 2.12 (s, 3H,
OCOCH3), 2.48 (br, 4H, NCH2), 5.27 (s, 1H, C17-␣H), 6.07 (d, 1H,
CH, J = 2.36 Hz), 7.20 (s, 4H). 13C NMR (150 MHz, CDCl3): ı 12.27,
12.32, 20.54, 21.18, 23.18, 28.01, 28.70, 30.95, 31.74, 34.77, 34.86,
35.88, 36.49, 37.37, 42.99, 45.21, 48.84, 51.86, 54.39, 64.33, 84.55,
122.36, 128.43, 129.47, 132.02, 136.08, 141.96, 171.12. Anal. Calcd.
for C32H44ClNO2: C 75.34, H 8.69, N 2.75; Found C 75.09, H 8.51, N
2.58. MS (m/z): 510.4[M+1]+.
White solid. Yield 91%. m.p. 161–163 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.79 (s, 3H, CH3-18), 2.20 (s, 3H,
OCOCH3), 2.51 (br, 4H, NCH2), 3.86 (s, 9H, OCH3), 5.35 (s, 1H, C17
-
␣H), 6.13 (d, 1H, CH, J = 2.20 Hz), 6.57 (s, 2H). 13C NMR (150 MHz,
CDCl3): ı 12.33, 12.39, 20.62, 21.24, 24.03, 24.82, 26.34, 28.78,
30.63, 30.93, 31.73, 34.84, 36.07, 36.50, 37.91, 42.98, 45.98, 48.87,
50.32, 54.36, 56.08, 60.92, 64.46, 84.60, 105.53, 123.58, 133.41,
136.80, 140.51, 152.95, 171.25. Anal. Calcd. for C36H53NO5: C
74.57, H 9.21, N 2.42; Found C 74.66, H 9.08, N 2.19. MS (m/z):
580.6[M+1]+.
2.6.19. 16-(4-chloro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-yl acetate (14e)
White solid. Yield 95%. m.p. 86–88 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.58 (s, 4H, NCH2), 3.73–3.75 (m, 4H, OCH2), 5.35 (s,
1H, C17-␣H), 6.16 (d, 1H, CH, J = 2.28 Hz), 7.29 (s, 4H). 13C NMR
(150 MHz, CDCl3): ı 11.58, 12.27, 12.36, 20.60, 21.17, 24.50, 28.76,
30.93, 31.10, 31.70, 34.85, 35.99, 37.64, 42.99, 45.66, 46.21, 48.81,
50.06, 54.34, 64.07, 67.36, 84.52, 122.40, 128.43, 129.47, 132.04,
136.06, 141.89, 171.12. Anal. Calcd. for C32H44ClNO3: C 73.05, H
8.43, N 2.66; Found C 73.33, H 8.51, N 2.47. MS (m/z): 526.6[M+1]+.
2.6.14. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-yl acetate (13d)
White solid. Yield 96%. m.p. 117–119 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.20 (s, 3H,
OCOCH3), 2.53 (br, 4H, NCH2), 3.86 (s, 9H, OCH3), 5.35 (s, 1H, C17
-
␣H), 6.13 (d, 1H, CH, J = 2.20 Hz), 6.57 (s, 2H). 13C NMR (150 MHz,
CDCl3): ı 12.31, 14.19, 20.55, 21.22, 23.16, 27.93, 28.70, 30.91,
31.74, 34.70, 34.84, 35.88, 36.51, 37.35, 42.97, 45.20, 48.90, 51.81,
54.38, 56.09, 60.38, 60.90, 64.30, 84.59, 105.56, 123.59, 133.40,
136.84, 140.50, 152.96, 171.21. Anal. Calcd. for C35H51NO5: C
74.30, H 9.09, N 2.48; Found C 74.08, H 8.89, N 2.57. MS (m/z):
566.7[M+1]+.
2.6.20. 16-(4-chloro-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-yl acetate (15e)
White solid. Yield 95%. m.p. 124–126 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.75 (s, 3H, CH3-19), 0.79 (s, 3H, CH3-18), 2.19 (s, 3H,
OCOCH3), 2.27 (s, 3H, Ph-CH3), 2.46–2.61 (m, 8H, NCH2), 5.35 (s,
1H, C17-␣H), 6.14 (d, 1H, CH, J = 2.24 Hz), 7.28 (s, 4H). 13C NMR
(150 MHz, CDCl3): ı 12.25, 12.36, 20.58, 21.14, 24.50, 28.74, 30.91,
31.23, 31.68, 34.82, 35.95, 37.72, 42.96, 45.76, 46.01, 48.78, 49.26,
54.33, 55.48, 63.75, 84.49, 122.35, 128.41, 129.45, 132.00, 136.06,
141.89, 171.02. Anal. Calcd. for C33H47ClN2O2: C 73.51, H 8.79, N
5.20; Found C 73.42, H 8.66, N 5.03. MS (m/z): 539.6[M+1]+.
2.6.15. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-yl acetate (14d)
White solid. Yield 93%. m.p. 176–178 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.80 (s, 3H, CH3-18), 2.20 (s, 3H,
OCOCH3), 2.56 (s, 4H, NCH2), 3.72–3.74 (m, 4H, OCH2), 3.86 (s,
9H, OCH3), 5.35 (s, 1H, C17-␣H), 6.13 (d, 1H, CH, J = 2.12 Hz), 6.57
(s, 2H). 13C NMR (150 MHz, CDCl3): ı 12.18, 12.31, 20.57, 21.17,
24.42, 28.72, 30.86, 31.08, 31.65, 34.78, 35.94, 36.45, 37.58, 42.92,
45.59, 46.14, 48.80, 50.01, 54.28, 56.02, 60.83, 63.99, 67.29, 84.50,
105.51, 123.57, 133.33, 136.79, 140.35, 152.91, 171.10. Anal. Calcd.
for C35H51NO6: C 72.26, H 8.84, N 2.41; Found C 72.09, H 8.77, N
2.56. MS (m/z): 582.6[M+1]+.
2.6.21. 16-(4-bromo-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-yl acetate (12f)
White solid. Yield 94%. m.p. 157–159 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.67 (s, 3H, CH3-19), 0.71 (s, 3H, CH3-18), 2.12 (s, 3H,
OCOCH3), 2.45 (s, 4H, NCH2), 5.25 (s, 1H, C17-␣H), 6.05 (d, 1H, CH,
J = 2.00 Hz), 7.14 (d, 2H, J = 8.44 Hz), 7.37 (d, 2H, J = 8.48 Hz). 13C NMR
(150 MHz, CDCl3): ı 12.28, 12.39, 20.61, 21.19, 23.99, 24.75, 26.21,
28.76, 30.53, 30.96, 31.71, 34.85, 36.05, 37.88, 42.99, 45.96, 48.80,
50.30, 54.34, 64.50, 84.55, 120.20, 122.42, 129.80, 131.38, 136.51,
142.17, 171.12. Anal. Calcd. for C33H46BrNO2: C 69.70, H 8.15, N
2.46; Found C 69.55, H 8.31, N 2.29. MS (m/z): 569.6[M+1]+.
2.6.16. 16-(3,4,5-trimethoxy-benzylidene)-3ˇ-(4-
methylpiperazinyl)-5˛-androstan-17ˇ-yl acetate (15d)
White solid. Yield 92%. m.p. 159–161 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.78 (s, 3H, CH3-19), 0.79 (s, 3H, CH3-18), 2.20 (s, 3H,
OCOCH3), 2.28 (s, 3H, Ph-CH3), 2.46–2.62 (m, 8H, NCH2), 3.86 (s, 9H,
OCH3), 5.35 (s, 1H, C17-␣H), 6.13 (d, 1H, CH, J = 2.12 Hz), 6.57 (s,
2H). 13C NMR (150 MHz, CDCl3): ı 12.31, 12.34, 20.59, 21.19, 24.39,
28.73, 30.88, 31.17, 31.67, 34.82, 35.95, 37.69, 42.95, 45.74, 45.90,
48.84, 49.13, 54.32, 55.31, 56.07, 60.87, 63.74, 84.55, 105.58, 123.58,
133.37, 136.85, 140.42, 152.94, 171.17. Anal. Calcd. for C36H54N2O5:
C 72.69, H 9.15, N 4.71; Found C 72.55, H 9.37, N 4.51. MS (m/z):
595.7[M+1]+.
2.6.22. 16-(4-bromo-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-yl acetate (13f)
White solid. Yield 94%. m.p. 184–186 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.76 (s, 3H, CH3-19), 0.82 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.57 (br, 4H, NCH2), 5.34 (s, 1H, C17-␣H), 6.14 (d, 1H, CH,
J = 2.04 Hz), 7.22 (d, 2H, J = 8.44 Hz), 7.45 (d, 2H, J = 8.48 Hz). 13C NMR
(150 MHz, CDCl3): ı 12.26, 12.32, 20.54, 21.17, 23.17, 27.93, 28.69,
30.96, 31.72, 34.69, 34.84, 35.87, 36.47, 37.35, 42.98, 45.20, 48.82,
51.82, 54.37, 64.31, 84.54, 120.19, 122.40, 129.80, 131.37, 136.50,
142.17, 171.08. Anal. Calcd. for C32H44BrNO2: C 69.30, H 8.00, N
2.53; Found C 69.06, H 7.83, N 2.41. MS (m/z): 554.5[M+1]+.
2.6.17. 16-(4-chloro-benzylidene)-3ˇ-piperidino-5˛-
androstan-17ˇ-yl acetate (12e)
White solid. Yield 92%. m.p. 99–101 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.80 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.51 (s, 4H, NCH2), 5.35 (s, 1H, C17-␣H), 6.16 (d, 1H, CH,
J = 2.28 Hz), 7.28 (s, 4H). 13C NMR (150 MHz, CDCl3): ı 11.62, 12.28,
12.40, 20.61, 21.18, 24.17, 24.89, 26.47, 28.79, 30.69, 30.95, 31.74,
34.87, 36.08, 37.95, 43.00, 46.02, 46.24, 48.83, 50.39, 54.39, 64.51,

720
H. Guo et al. / Steroids 76 (2011) 709–723
2.6.23. 16-(4-bromo-benzylidene)-3ˇ-morpholino-5˛-
2.6.28. 16-(4-fluoro-benzylidene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-yl acetate (14f)
androstan-17ˇ-yl acetate (15g)
White solid. Yield 90%. m.p. 184–186 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.77 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.21 (s, 3H,
OCOCH3), 2.58 (s, 4H, NCH2), 3.72–3.76 (m, 4H, OCH2), 5.35 (s,
1H, C17-␣H), 6.15 (d, 1H, CH, J = 2.04 Hz), 7.23 (d, 2H, J = 8.44 Hz),
7.46 (d, 2H, J = 8.48 Hz). 13C NMR (150 MHz, CDCl3): ı 12.26, 12.37,
20.65, 21.21, 23.97, 24.68, 26.22, 28.77, 30.55, 30.98, 31.70, 34.83,
36.03, 37.90, 42.92, 45.95, 48.84, 50.31, 54.35, 64.53, 84.60, 120.24,
122.48, 129.81, 131.37, 136.53, 142.20, 171.09. Anal. Calcd. for
C32H44BrNO3: C 67.36, H 7.77, N 2.45; Found C 67.21, H 7.59, N
2.28. MS (m/z): 571.6[M+1]+.
White solid. Yield 94%. m.p. 178–180 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.75 (s, 3H, CH3-19), 0.79 (s, 3H, CH3-18), 2.19 (s, 3H,
OCOCH3), 2.27 (s, 3H, Ph-CH3), 2.46–2.61 (m, 8H, NCH2), 5.34 (s, 1H,
C17-␣H), 6.15 (d, 1H, CH, J = 2.00 Hz), 7.00 (t, 2H), 7.30 (q, 2H). 13
C
NMR (150 MHz, CDCl3): ı 12.24, 12.35, 20.58, 21.12, 24.52, 28.74,
30.79, 31.25, 31.68, 34.83, 35.94, 37.72, 42.94, 45.76, 46.03, 48.80,
49.28, 54.33, 55.51, 63.74, 84.47, 115.01, 115.23, 122.37, 129.69,
129.77, 133.72, 133.75, 140.58, 160.04, 162.49, 171.00. Anal. Calcd.
for C33H47FN2O2: C 75.82, H 9.06, N 5.36; Found C 75.69, H 8.89, N
5.18. MS (m/z): 523.5[M+1]+.
2.6.24. 16-(4-bromo-benzylidene)-3ˇ-(4-
methylpiperazinyl)-5˛-androstan-17ˇ-yl acetate (15f)
2.6.29. 16-(furan-2-ylmethylene)-3ˇ-piperidino-5˛-
androstan-17ˇ-yl acetate (12h)
White solid. Yield 98%. m.p. 172–174 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.74 (s, 3H, CH3-19), 0.79 (s, 3H, CH3-18), 2.19 (s, 3H,
OCOCH3), 2.27 (s, 3H, Ph-CH3), 2.46–2.61 (m, 8H, NCH2), 5.32 (s,
1H, C17-␣H), 6.13 (d, 1H, CH, J = 2.16 Hz), 7.20 (d, 2H, J = 8.52 Hz),
7.43 (d, 2H, J = 8.44 Hz). 13C NMR (150 MHz, CDCl3): ı 12.26, 12.37,
20.59, 21.15, 24.52, 28.75, 30.93, 31.25, 31.69, 34.84, 35.97, 37.74,
42.97, 45.78, 46.04, 48.79, 49.30, 54.35, 55.52, 63.77, 84.52, 120.18,
122.42, 129.79, 131.36, 136.50, 142.14, 171.04. Anal. Calcd. for
White solid. Yield 94%. m.p. 184–186 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.74 (s, 3H, CH3-19), 0.80 (s, 3H, CH3-18), 2.19 (s, 3H,
OCOCH3), 2.53 (s, 4H, NCH2), 5.34 (s, 1H, C17-␣H), 6.13 (d, 1H, CH,
J = 2.32 Hz), 6.22 (d, 1H, J = 2.32 Hz), 6.41 (t, 1H), 7.38 (s, 1H). 13C
NMR (150 MHz, CDCl3): ı 12.24, 12.41, 20.65, 21.13, 24.13, 24.88,
26.45, 28.83, 30.71, 31.08, 31.74, 34.89, 36.08, 37.96, 43.36, 46.05,
48.64, 50.37, 54.43, 64.52, 84.27, 107.83, 111.26, 112.16, 140.01,
141.29, 153.42, 171.08. Anal. Calcd. for C31H45NO3: C 77.62, H 9.46,
N 2.92; Found C 77.47, H 9.33, N 3.09. MS (m/z): 480.4[M+1]+.
C
33H47BrN2O2: C 67.91, H 8.12, N 4.80; Found C 67.88, H 8.29, N
4.61. MS (m/z): 583.6[M+1]+.
2.6.25. 16-(4-fluoro-benzylidene)-3ˇ-piperidino-5˛-
2.6.30. 16-(furan-2-ylmethylene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-yl acetate (12g)
androstan-17ˇ-yl acetate (13h)
White solid. Yield 90%. m.p. 165–167 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.68 (s, 3H, CH3-19), 0.71 (s, 3H, CH3-18), 2.12 (s, 3H,
OCOCH3), 2.47 (s, 4H, NCH2), 5.26 (s, 1H, C17-␣H), 6.08 (d, 1H, CH,
J = 1.96 Hz), 6.93 (t, 2H), 7.23 (q, 2H). 13C NMR (150 MHz, CDCl3): ı
11.35, 12.26, 12.37, 20.61, 21.16, 23.96, 24.75, 26.20, 28.76, 30.51,
30.83, 31.70, 34.85, 36.03, 37.87, 42.97, 45.95, 46.07, 48.82, 50.25,
54.35, 64.44, 84.52, 115.04, 115.25, 122.38, 129.71, 129.79, 133.74,
133.77, 140.63, 160.07, 162.52, 171.11. Anal. Calcd. for C33H46FNO2:
C 78.06, H 9.13, N 2.76; Found C 78.31, H 9.01, N 2.55. MS (m/z):
508.5[M+1]+.
White solid. Yield 94%. m.p. 155–157 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.74 (s, 3H, CH3-19), 0.83 (s, 3H, CH3-18), 2.19 (s, 3H,
OCOCH3), 2.57 (br, 4H, NCH2), 5.35 (s, 1H, C17-␣H), 6.13 (d, 1H, CH,
J = 2.26 Hz), 6.22 (d, 1H, J = 2.30 Hz), 6.41 (t, 1H), 7.38 (s, 1H). 13C
NMR (150 MHz, CDCl3): ı 12.22, 12.32, 20.57, 21.12, 23.17, 27.97,
28.73, 31.06, 31.72, 34.75, 34.86, 35.87, 36.51, 37.37, 43.34, 45.24,
48.65, 51.82, 54.42, 64.32, 84.24, 107.82, 111.24, 112.14, 139.96,
141.26, 153.39, 171.03. Anal. Calcd. for C30H43NO3: C 77.38, H 9.31,
N 3.01; Found C 77.51, H 9.09, N 3.12. MS (m/z): 466.4[M+1]+.
2.6.31. 16-(furan-2-ylmethylene)-3ˇ-morpholino-5˛-
androstan-17ˇ-yl acetate (14h)
2.6.26. 16-(4-fluoro-benzylidene)-3ˇ-pyrrolidino-5˛-
androstan-17ˇ-yl acetate (13g)
White solid. Yield 94%. m.p. 194–196 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.74 (s, 3H, CH3-19), 0.81 (s, 3H, CH3-18), 2.19 (s, 3H,
OCOCH3), 2.58 (s, 4H, NCH2), 3.70–3.73 (m, 4H, OCH2), 5.35 (s, 1H,
C17-␣H), 6.13 (d, 1H, CH, J = 2.36 Hz), 6.22 (d, 1H, J = 3.32 Hz), 6.41
(t, 1H), 7.38 (s, 1H). 13C NMR (150 MHz, CDCl3): ı 12.23, 12.37,
20.64, 21.13, 24.50, 28.80, 31.06, 31.12, 31.70, 34.86, 36.00, 37.65,
43.35, 45.69, 48.62, 50.06, 54.38, 64.08, 67.31, 84.23, 107.85, 111.26,
112.17, 139.91, 141.30, 153.39, 171.07. Anal. Calcd. for C30H43NO4:
C 74.81, H 9.00, N 2.91; Found C 74.66, H 8.83, N 3.12. MS (m/z):
482.4[M+1]+.
White solid. Yield 93%. m.p. 107–109 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.68 (s, 3H, CH3-19), 0.75 (s, 3H, CH3-18), 2.12 (s, 3H,
OCOCH3), 2.72 (br, 4H, NCH2), 5.26 (s, 1H, C17-␣H), 6.08 (d, 1H, CH,
J = 2.00 Hz), 6.93 (t, 2H), 7.23 (q, 2H). 13C NMR (150 MHz, CDCl3): ı
10.32, 12.21, 12.26, 20.56, 21.18, 23.23, 26.36, 28.54, 30.81, 31.63,
33.04, 34.80, 35.79, 36.45, 37.13, 42.96, 45.20, 48.82, 51.13, 54.22,
64.07, 84.51, 115.05, 115.26, 122.43, 129.71, 129.79, 133.72, 133.75,
140.58, 160.08, 162.54, 171.20. Anal. Calcd. for C32H44FNO2: C
77.85, H 8.98, N 2.84; Found C 77.69, H 8.79, N 2.77. MS (m/z):
494.5[M+1]+.
2.6.27. 16-(4-fluoro-benzylidene)-3ˇ-morpholino-5˛-
androstan-17ˇ-yl acetate (14g)
2.6.32. 16-(furan-2-ylmethylene)-3ˇ-(4-methylpiperazinyl)-5˛-
androstan-17ˇ-yl acetate (15h)
White solid. Yield 94%. m.p. 141–143 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.68 (s, 3H, CH3-19), 0.72 (s, 3H, CH3-18), 2.12 (s, 3H,
OCOCH3), 2.57 (s, 4H, NCH2), 3.68–3.71 (m, 4H, OCH2), 5.27 (s, 1H,
White solid. Yield 94%. m.p. 145–147 ◦C; 1H NMR (400 MHz,
CDCl3): ı 0.72 (s, 3H, CH3-19), 0.79 (s, 3H, CH3-18), 2.17 (s, 3H,
OCOCH3), 2.28 (s, 3H, Ph-CH3), 2.46–2.61 (m, 8H, NCH2), 5.32 (s,
1H, C17-␣H), 6.11 (d, 1H, CH, J = 2.28 Hz), 6.20 (d, 1H, J = 3.28 Hz),
6.38 (t, 1H), 7.36 (s, 1H). 13C NMR (150 MHz, CDCl3): ı 12.18, 12.34,
20.58, 21.02, 24.49, 28.76, 31.00, 31.26, 31.64, 34.80, 35.91, 37.70,
43.27, 45.74, 46.02, 48.56, 49.26, 54.34, 55.51, 63.71, 84.12, 107.80,
111.21, 112.14, 139.83, 141.21, 153.33, 170.81. Anal. Calcd. for
C17-␣H), 6.08 (d, 1H, CH, J = 2.00 Hz), 6.93 (t, 2H), 7.23 (q, 2H). 13
C
NMR (150 MHz, CDCl3): ı 12.28, 20.60, 21.18, 23.80, 28.68, 30.39,
30.81, 31.64, 34.83, 35.92, 37.47, 42.97, 45.56, 48.81, 49.55, 54.25,
64.15, 66.61, 84.51, 115.06, 115.27, 122.45, 129.72, 129.79, 133.72,
133.76, 140.54, 160.10, 162.55, 171.18. Anal. Calcd. for C32H44FNO3:
C 75.41, H 8.70, N 2.75; Found C 75.27, H 8.58, N 2.91. MS (m/z):
510.4[M+1]+.
C
31H46N2O3: C 75.26, H 9.37, N 5.66; Found C 75.08, H 9.51, N 5.44.
MS (m/z): 495.4[M+1]+.

H. Guo et al. / Steroids 76 (2011) 709–723
721
2.7. Biology
2.7.1. Cell lines and culture conditions
Human cancer cell lines SW480 [25], A549 [26], HepG2 [27] and
HeLa [28,29] used in this work, were purchased from China Cen-
tre for Type Culture Collection (Wuhan, China). SW480, A549 and
HepG2 cells were grown in RPMI 1640 supplemented with 10%
fetal bovine serum (FBS), 10 U penicillin and 100 ␮g/ml strepto-
mycin at 37 ◦C with 5% CO2 in a humidified atmosphere. HeLa cells
were grown in Dulbecco’s modified Eagle’s medium (DMEM) sup-
planted with FCS and aitibiotics as described above for RPMI 1640.
Fresh medium was given every second day and on the day before
the experiments were done. Cells were passaged at preconfluent
densities, using a solution containing 0.05% trypsin and 0.5 mM
EDTA.
Fig. 2. X-ray crystal structure of 8e.
to CCDC, 12 Union Road, Cambridge CB2 1EZ, UK [fax (+44) 1223
336033, e-mail: deposit@ccdc.cam.ac.uk].
3.2. Cytotoxic activity
2.7.2. Cell viability assay (MTT)
The anticancer activity in vitro was measured using the MTT
assay. The assay was carried out according to previous study
[30,31]. Exponentially growing cells were harvested and plated
in 96-well plates at a concentration of 1 × 104 cells/well. After
24 h incubation at 37 ◦C under a humidified 5% CO2 to allow cell
attachment, the cells in the wells were respectively treated with
target compounds at various concentrations for 48 h. The concen-
tration of DMSO was always kept below 1.25%, which was found
to be non-toxic to the cells. A solution of 3-(4,5-dimethylthizao1-
2-y1)-2,5-diphenyltetrazolium bromide (MTT), was prepared at
5 mg/ml in phosphate buffered saline (PBS; 1.5 mM KH2PO4,
6.5 mM Na2HPO4, 137 mM NaCI, 2.7 mM KCI; pH 7.4). Of this solu-
tion 20 ␮l was added to each well. After incubation for 4 h at 37 ◦C
in a humidified incubator with 5% CO2, the medium/MTT mixtures
were removed, and the formazan crystals formed by the mitochon-
drial dehydrogenase activity of vital cells were dissolved in 100 ␮l
of DMSO per well. The absorbance of the wells was read with a
microplate reader (Bio-Rad Instruments) at 570 nm. Effects of the
drug cell viability were calculated using cell treated with DMSO as
control.
The newly synthesized compounds 12a–h, 13a–h, 14a–h,
15a–h were evaluated for their anticancer activity towards human
tumor cell lines derived from various human cancer types: SW480
(human colon adenocarcinoma cells), HeLa (human cervical cancer
cells), A549 (human lung carcinoma cells), HepG2 (human hepatic
carcinoma cells). In vitro evaluation of anticancer activity of
the synthesized compounds was carried out using the 3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)
assay. Anticancer potency of the compounds was indicated by IC50
values that were calculated by linear regression analysis of the
concentration–response curves obtained for each compound. Data
were reported in Table 1.
Table 1
Cytotoxic activity data for compounds 12a–h, 13a–h, 14a–h, 15a–h against SW480,
A549, HepG2, and HeLa.
a
Compounds
IC50 (␮mol l−1
)
SW480
A549
6.18
13.88
38.97
>50
9.03
HepG2
HeLa
20.21
28.51
>50
40.58
17.26
23.89
12a
13a
14a
15a
12b
13b
14b
15b
12c
13c
14c
15c
12d
13d
14d
15d
12e
13e
14e
15e
12f
21.68
15.54
40.56
>50
12.86
30.18
>50
>50
>50
>50
>50
12.33
22.47
>50
2.7.3. Data analysis
Cell survival was calculated using the formula: Survival
(%) = [(absorbance of treated cells − absorbance of culture
medium)/(absorbance of untreated cells − absorbance of culture
medium)] × 100 [32,33]. The experiment was done in triplicate
and the inhibitory concentration (IC) values were calculated from
a dose response curve. IC50 is the concentration in ␮M required
for 50% inhibition of cell growth as compared to that of untreated
control. IC50 values were determined from the linear portion of
the curve by calculating the concentration of agent that reduced
absorbance in treated cells, compared to control cells, by 50%.
Evaluation is based on mean values from three independent
experiments, each comprising at least six microcultures per
concentration level.
>50
6.54
10.55
39.48
>50
38.21
>50
17.76
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
11.56
49.18
>50
>50
>50
13.37
>50
38.77
>50
>50
>50
>50
38.83
>50
23.10
19.89
44.51
41.73
31.42
>50
>50
>50
>50
>50
21.58
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
4.07
6.14
16.58
>50
>50
>50
>50
>50
>50
41.03
29.86
>50
>50
35.46
19.20
37.52
>50
15.53
30.19
42.41
3. Results and discussion
13f
14f
15f
3.1. Chemistry
>50
>50
12g
13g
14g
15g
12h
13h
14h
15h
17.92
6.45
11.71
>50
21.19
>50
6.78
23.61
33.54
>50
30.79
15.52
37.71
>50
We obtained an X-ray crystal structure of 8e that is presented
in Fig. 2. The configuration at C-16 has been assigned E which was
same with the earlier report [34,35]. This structure confirms the ␤-
stereochemistry of the heterocycles at C-3 and the hydroxyl group
at C-17. After the acetylation of 17␤-hydroxy derivatives, the ␤
configuration of acetoxy group at C-17 was maintained. Crystal-
lographic data for 8e have been deposited with the Cambridge
Crystallographic Data Centre with the deposition number CCDC
796339. These data can be obtained free of charge on application
13.83
23.18
27.23
>50
>50
>50
>50
Exponentially growing cells were treated with the compounds at different concen-
trations for 48 h. Cell-growth inhibition was analysed by the MTT assay.
a
IC50 is the concentration of compound that inhibits 50% of cell growth.

722
H. Guo et al. / Steroids 76 (2011) 709–723
In conclusion, we have described a facile synthesis of novel
promising anti-cancer steroid derivatives. This study may provide
valuable information for the further design and development of
more potent anticancer agents.
Acknowledgments
This work was financially supported by the Important National
Science & Technology Specific Projects (No. 2009ZX09301-014-1)
and the Fundamental Research Funds for the Central Universities.
Fig. 3. General SAR of compounds for anticancer activity in vitro based on the IC50
results on the four cancer cells lines.
As shown in Table 1, compounds 12–15a, 12–14b, 12–14d,
12–13e, 12–13f, 12–15g, 12–15h exhibited different levels of
anticancer properties; namely, the introduction of methyl and
methoxy at the para position of the phenyl ring lead to potent
anticancer activity. For A549, among the 12–15a, 12–15b bear-
ing methyl and methoxy group, the IC50 values obtained are
6.18 ␮M, 13.88 ␮M, 38.97 ␮M, >50 ␮M, 9.03 ␮M, 17.76 ␮M, >50 ␮M
and >50 ␮M, respectively. It was noteworthy that compound 12b
which has methoxy at the 4-position of the phenyl ring exhib-
ited potent activity (12b for A549 9.03 ␮M, for HepG2 6.54 ␮M).
Compared to 12–15b, the inclusion of more methoxy groups
(12–14d) to the phenyl ring enabled maintenance of anticancer
activity. On the other hand, substitution with a strong electron-
withdrawing group like nitro (12–15c) at the para position of
the phenyl ring resulted in a complete loss of anticancer activity
(IC50 > 50 ␮M). Comparison of the results obtained from p-methyl,
p-methoxy, and p-nitro derivatives indicated that the electron-
donating groups were relatively beneficial for anticancer activity.
However, there was an exception, the IC50 value of compound
12c against HepG2 was 38.21 ␮M. When the alkyl groups were
replaced by a halogen (12–15e, 12–15f) such as chlorine or bromine
at the para position of the phenyl ring, decreased cytotoxicity
was observed (>50 ␮M). Among the tested compounds, it was
determined that 12-15 g exhibited potent activity against all four
cancer cell lines. The p-fluorophenyl derivative 12g exhibited
remarkable anticancer activity against A549 (4.07 ␮M) and HepG2
(6.78 ␮M). When the phenyl rings were replaced by a furan ring,
12–15h exhibited moderate activity against the A549 from 13.83
to 27.23 ␮M, suggesting that the phenyl ring at the C-16 posi-
tion may be unnecessary for the activity. Furthermore, it appears
that an appropriately substituted group of the phenyl ring at
C-16 could serve as a promising launch point for the further
design of this type of steroidal anticancer agent. A similar ten-
dency was observed concerning the SW480, HepG2, and HeLa cell
lines.
Most derivatives with a saturated heterocyclic ring moiety at C-3
position resulted in high activity, especially piperidine. For HepG2,
as demonstrated in Table 1, the most highly active compound was
12g with a piperidine moiety, having a IC50 value of 6.78 ␮M, fol-
lowed by 13g with a pyrrolidine moiety, exhibiting a IC50 value
of 23.61 ␮M, the derivative 14g with a morpholine moiety, which
exhibited a lower activity (33.54 ␮M), and finally the derivative 15g
with a methyl piperazine moiety exhibited no activity (>50 ␮M).
General SAR of compounds for anticancer activity in vitro based on
the IC50 results on the four cancer cells lines is presened schemat-
ically in Fig. 3.
In the present study, 32 novel 16E-arylidene-3␤-aminosteroid
derivatives were synthesized and evaluated for their anticancer
activity against four human cancer cell lines (SW480, A549, HepG2,
and HeLa) in vitro. Most of the compounds showed moderate to
good anticancer activity. Among the target derivatives, compounds
12a, 12b, 12g and 13g were found to have significant activity against
all four cancer cell lines. In particular, compounds 12b and 12g were
the most promising derivatives.
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