146
M. Kahnt et al. / European Journal of Medicinal Chemistry 159 (2018) 143e148
ꢀ
ꢀ
1
55e159 C; [
a
]
D
¼ þ27.26 (c 0.310, MeOH); R
f
¼ 0.21 (CHCl
3
/
acetone, 2:1); IR (ATR):
516m, 1448m, 1375m, 1252w, 1048s, 961m, 698s cm ; UV-Vis
n
¼ 3366br w, 2917m, 2859w, 1638m,
ꢁ
1
1
1
(CHCl
3
):
l
max (log ε) ¼ 259 nm (2.45); H NMR (400 MHz, CDCl
3
):
d
¼ 7.35e7.30 (m, 2H, H
m p
-Ph), 7.30e7.25 (m, 1H, H -Ph), 7.25e7.21
(
m, 2H, H
H), 4.54 (dd, J ¼ 14.6, 5.9 Hz,1H, CH
CH
Ph), 3.75 (ddd, J ¼ 10.7, 10.5, 4.4 Hz, 1H, 2-H), 3.61 (d, J ¼ 10.7 Hz,
H, 23-H
), 3.42 (d, J ¼ 9.4 Hz, 1H, 3-H), 3.37 (d, J ¼ 10.6 Hz, 1H, 23-
), 2.05e1.22 (m,17H,16-H ,1-H ,11-H , 22-H ,18-H,11-H ,16-H
5-H , 9-H, 21-H , 22-H , 7-H , 19-H, 6-H , 6-H , 21-H , 7-H ), 1.08
s, 3H, 27-H), 1.07e0.99 (m, 2H, 5-H, 15-H ), 0.98 (s, 3H, 25-H), 0.94
br s, 3H, 30-H), 0.97e0.89 (m, 2H, 20-H, 1-H
), 0.84 (d, J ¼ 5.6 Hz,
H, 29-H), 0.83 (s, 3H, 24-H), 0.69 (s, 3H, 26-H) ppm; C NMR
100 MHz, CDCl ): -Ph),
¼ 178.2 (C-28), 140.0 (C-13), 138.3 (C
-Ph), 128.0 (C -Ph), 127.6 (C -Ph), 125.7 (C-12), 80.0 (C-3),
9.7 (C-23), 68.7 (C-2), 54.1 (C-18), 48.8 (C-5), 47.9 (C-17), 47.5 (C-9),
6.3 (C-1), 43.9 (CH Ph), 42.7 (C-14, C-4), 39.9 (C-19), 39.7 (C-8),
9.2 (C-20), 38.1 (C-10), 37.4 (C-22), 32.5 (C-7), 31.0 (C-21), 28.0 (C-
5), 25.0 (C-16), 23.5 (C-27), 23.5 (C-11), 21.3 (C-30), 18.4 (C-6), 17.3
C-29), 17.3 (C-25), 17.2 (C-26), 13.1 (C-24) ppm; MS (ESI, MeOH): m/
o
-Ph), 6.17 (t, J ¼ 5.3 Hz, 1H, NH), 5.21 (t, J ¼ 3.4 Hz, 1H, 12-
a
Ph), 4.15 (dd, J ¼ 14.6, 4.3 Hz,1H,
b
1
H
1
(
(
a
b
a
a
a
a
b
b
,
a
a
b
a
a
b
b
b
b
b
13
3
(
3
d
i
1
6
4
3
28.8 (C
m
o
p
2
1
(
þ
þ
z ¼ 578.3 (100%, [MþH] ), 600.4 (10%, [MþNa] ), 1155.3 (34%,
þ
þ
[
2MþH] ), 1177.5 (45%, [2MþNa] ); analysis calcd for C37
H55NO
4
Fig. 2. Comparison of cytotoxicity of EM2 and compounds 1e3, 8 and 11 for A2780
tumor cell line vs. non-malignant mouse fibroblasts NIH 3T3 (EC50 from SRB assays).
Selectivity index (SI) is defined as: SI ¼ EC50 (NIH 3T3)/EC50 (A2780).
(577.85): C 76.91, H 9.59, N 2.42; found: C 76.70, H 9.82, N 2.37.
4.1.4. 2
a,3b,23-Tris(acetyloxy)-N-piperazinyl-urs-12-en-28-amide
(
8)
1
28.8 (C
0.0 (C-2), 65.4 (C-23), 54.1 (C-18), 47.9 (C-17), 47.7 (C-5), 47.6 (C-9),
3.9 (C-1), 43.8 (CH Ph), 42.6 (C-14), 42.1 (C-4), 39.9 (C-19), 39.7 (C-
), 39.2 (C-20), 37.9 (C-10), 37.4 (C-22), 32.5 (C-7), 31.0 (C-21), 27.9
m o p
-Ph), 128.1 (C -Ph), 127.6 (C -Ph), 125.3 (C-12), 74.9 (C-3),
To an ice-cold solution of 1 (200 mg, 0.32 mmol) in dry DCM
15 mL), triethylamine (0.01 mL, 0.07 mmol), DMF (0.005 mL,
.07 mmol) and oxalyl chloride (0.14 mL, 1.60 mmoL) were added.
7
4
8
(
0
2
After 2 h of stirring at room temperature, the solution was
concentrated under diminished pressure. Piperazine (83 mg,
(
2
1
7
C-15), 24.9 (C-16), 23.5 (C-11), 23.3 (C-27), 21.3 (C-30), 21.2 (Ac),
1.0 (Ac), 20.9 (Ac), 18.0 (C-6), 17.3 (C-29), 17.2 (C-25), 17.1 (C-26),
0.96 mmol) was dissolved in dry DCM (15 mL); triethylamine
þ
4.1 (C-24) ppm; MS (ESI, MeOH): m/z ¼ 704.4 (100%, [MþH] ),
(0.07 mL, 0.48 mmol), DMAP (4 mg, 0.04 mmol) and the acid chlo-
þ
þ
26.5 (48%, [MþNa] ), 1407.2 (26%, [2MþH] ), 1429.3 (32%,
ride were added. The solution was stirred for 1 h at room temper-
ature. The solvent was removed under diminished pressure, and
the residue was subjected to column chromatography (SiO , chlo-
2
þ
[
2MþNa] ); analysis calcd for C43
7
H61NO (703.96): C 73.37, H 8.73,
N 1.99; found: C 73.21, H 8.95, N 1.75.
roform/methanol, 9:1) to afforded 8 (190 mg, 86%) as a colorless
ꢀ
ꢀ
4
.1.3. 2
a
,3
b
,23-Trihydroxy-N-(phenylmethyl)-urs-12-en-28-amide
solid; m.p. 157e160 C; [
(SiO
a
]
D
¼ 17.48 (c ¼ 0.27, CHCl
3
); R
f
¼ 0.32
(3)
2
, chloroform/methanol, 9:1); IR (KBr): v ¼ 3444s, 2972m,
To a solution of 2 (232 mg, 0.33 mmol) in methanol (10 mL) a
2948m, 2870m, 1746vs, 1630m, 1458m, 1370m, 1252vs, 1236vs,
ꢁ
1 1
solution of potassium hydroxide (92 mg, 1.65 mmol) in methanol
1044s cm ; H NMR (400 MHz, CDCl
1H, 12-H), 5.16 (ddd, J ¼ 11.1, 11.1, 4.6 Hz, 1H, 2-H), 5.07 (d,
J ¼ 10.3 Hz, 1H, 3-H), 3.84 (d, J ¼ 11.8 Hz, 1H, 23-H ), 3.64e3.50 (m,
5H, 23-H þ 38-H ), 2.85e2.77 (m, 4H, 37-H þ 37-H ),
þ 38-H
2.44 (d, J ¼ 9.2 Hz, 1H, 18-H), 2.08 (s, 3H, Ac), 2.08e2.03 (m, 1H, 1-
), 2.01 (s, 3H, Ac), 1.97 (s, 3H, Ac), 1.95e1.89 (m, 4H, 16-H
þ 16-
3
):
d
¼ 5.21 (dd, J ¼ 3.5, 3.5 Hz,
ꢀ
(
2 mL) was added. The mixture was stirred at 25 C for 20 h. After
completion of the reaction (as indicated by TLC), the solution was
concentrated under reduced pressure, and the residue was
precipitated in water (20 mL). Filtration and drying (CaCl
a
b
a
b
a
b
2
, 24 h)
afforded compound 3 (172 mg, 90%) as a colorless solid; m.p.
H
a
a
Fig. 3. Representative results of the SRB assay; dose-response curves of compound 11 for cell lines HT29 (colorectal carcinoma) and 518A2 (melanoma).