Journal of Medicinal Chemistry
Article
Table 4. EC50 Values (μM) of 13 and 14 with Human Melanoma Cells
compd primary melanoma wm3211 cell line metastatic melanoma A375 cell line primary fibroblast cells primary keratinocytes immortalized melanocytes
a
b
13
14
5.6
2.7
3.4
1.3
1.4
7.9
ND
cannot calculate
cannot calculate
cannot calculate
a
b
Not done. Toxic even at lower concentration and no dose−activity curve occurred.
extracted with EtOAc, and washed with water and brine. The organic
layer was concentrated in vacuo. The residue was purified by column
chromatography to yield 21 (yield 87%). Intermediate 24 was
synthesized by general procedures A and D using 21 as the starting
material (yield 46%). A solution of 24 (0.5 mmol) in MeOH (20 mL)
was treated with 10% Pd/C (15 mg). The reaction mixture was stirred
at room temperature under a hydrogen atmosphere for 24 h. The
catalyst was removed by filtration through Celite, and the resulting
solution was concentrated in vacuo. The crude material was purified by
2H), 4.28 (t, J = 6.0 Hz, 4H), 2.33 (p, J = 6.0 Hz, 2H). 13C NMR (125
MHz, CD3OD): δ 161.77, 158.79, 146.61, 134.93, 134.51, 132.32,
129.83, 118.43, 115.89, 112.62, 65.88, 30.21. LC-TOF (M + H+) calcd
for C25H25N4O2S2 477.1413, found 477.1411.
N-(3-(2-((3-(Thiophene-2-carboximidamido)benzyl)amino)-
ethyl)phenyl)thiophene-2-carboximidamide (13). 13 was syn-
thesized by general procedures B, C, A, D, and E using 3-
nitrobenzaldehyde and 2-(3-nitrophenyl)ethanamine as the starting
1
materials. H NMR (500 MHz, CD3OD): δ 7.65 (t, J = 5.0 Hz, 2H),
1
7.56 (dt, J = 5.0, 2.0 Hz, 2H), 7.38−7.27 (m, 2H), 7.17−7.09 (m, 2H),
7.06 (dt, J = 7.5, 1.5 Hz, 1H), 7.01−6.94 (m, 2H), 6.91 (dt, J = 7.5, 1.5
Hz, 1H), 6.88−6.82 (m, 2H), 3.79 (s, 2H), 2.96−2.81 (m, 4H). 13C
NMR (125 MHz, CD3OD): δ 161.52, 153.94, 150.23, 142.53, 141.90,
141.06, 130.73, 130.67, 129.87, 129.83, 128.49, 128.39, 125.03, 124.76,
123.95, 123.69, 122.60, 121.58, 54.30, 51.28, 36.59. LC-TOF (M +
H+) calcd for C25H26N5S2 460.1624, found 460.1618.
column chromatography to yield 9 (yield 91%). H NMR (400 MHz,
CD3OD): δ 8.17−8.02 (m, 4H), 7.56−7.51 (t, J = 7.5 Hz, 2H), 7.45−
7.38 (m, 6H), 7.36−7.30 (d, J = 7.5 Hz, 2H), 3.10 (s, 4H). 13C NMR
(100 MHz, CD3OD): δ 156.90, 143.97, 136.57, 133.15, 132.73,
130.64, 129.92, 128.32, 128.26, 125.00, 122.43, 36.92. LC-TOF (M +
H+) calcd for C24H23N4S2 431.1359, found 431.1359.
N-(3-(1-Hydroxy-2-(3-(thiophene-2-carboximidamido)-
phenoxy)ethyl)phenyl)thiophene-2-carboximidamide (10). To
a solution of 3-nitrophenol (2 mmol) in DMF (20 mL) was added
K2CO3 (2.2 mmol) and 2-bromo-1-(3-nitrophenyl)ethanone (2.2
mmol). The suspension was stirred for 6 h at room temperature. The
reaction mixture was then diluted with H2O (50 mL), extracted with
EtOAc, and washed with water and brine. The organic layer was dried
over Na2SO4 and concentrated in vacuo. The residue was purified by
column chromatography to yield 24 (yield 73%). To a solution of 24
(1 mmol) in THF (10 mL) was added NaBH4 (1.2 mmol) in three
portions. The reaction mixture was stirred at room temperature for 8 h
under a N2 atmosphere. The reaction mixture was quenched with H2O
(20 mL), extracted with CH2Cl2, and washed with water and brine.
The organic layer was dried over Na2SO4 and concentrated in vacuo.
The residue was purified by column chromatography to yield 25 (yield
80%). 10 was synthesized by general procedures A and D using 25 as
N-(4-(2-((3-(Thiophene-2-carboximidamido)benzyl)amino)-
ethyl)phenyl)thiophene-2-carboximidamide (14). 14 was syn-
thesized by the same procedures as those to prepare 13 using 2-(4-
nitrophenyl)ethanamine as the starting material. 1H NMR (500 MHz,
CD3OD): δ 7.65−7.58 (m, 2H), 7.58−7.51 (m, 2H), 7.33 (t, J = 7.5
Hz, 1H), 7.24−7.17 (m, 2H), 7.13−7.07 (m, 2H), 7.04 (dt, J = 7.5, 2.0
Hz, 1H), 6.95 (t, J = 2.0 Hz, 1H), 6.93−6.85 (m, 3H), 3.84−3.70 (m,
2H), 2.93−2.74 (m, 4H). 13C NMR (125 MHz, CD3OD): δ 153.93,
141.90, 141.03, 136.14, 130.99, 130.86, 130.66, 130.30, 129.86, 129.80,
128.50, 128.44, 128.37, 124.76, 123.80, 123.66, 123.60, 122.61, 54.27,
51.49, 36.09. LC-TOF (M + H+) calcd for C25H26N5S2 460.1624,
found 460.1620.
N-(4-(2-(Ethyl(3-(thiophene-2-carboximidamido)benzyl)-
amino)ethyl)phenyl)thiophene-2-carboximidamide (15). 35
was synthesized by general procedure B using 3-nitrobenzaldehyde
and 2-(4-nitrophenyl)ethanamine as the starting materials. To a
solution of 35 (1 mmol) and acetaldehyde (2 equiv) in THF (10 mL)
was added NaBH4 (1.2 mmol) in three portions. The reaction mixture
was stirred at room temperature for 8 h under a N2 atmosphere. The
reaction mixture was then quenched with H2O (20 mL), extracted
with CH2Cl2, and washed with water and brine. The organic layer was
dried over Na2SO4 and concentrated in vacuo. The residue was
purified by column chromatography to yield 39 (yield 64%). 15 was
synthesized by general procedures A and D using 39 as the starting
material. 1H NMR (500 MHz, CD3OD): δ 7.69−7.60 (m, 2H), 7.60−
7.54 (m, 2H), 7.36 (t, J = 7.5 Hz, 1H), 7.24−7.17 (m, 2H), 7.14−7.09
(m, 3H), 7.05−6.99 (m, 1H), 6.96−6.87 (m, 3H), 3.72 (s, 2H), 2.88−
2.73 (m, 4H), 2.72−2.64 (q, J = 7.0 Hz, 2H), 1.16 (t, J = 7.0 Hz, 3H).
13C NMR (125 MHz, CD3OD): δ 150.04, 141.15, 141.01, 136.83,
1
the starting material (yield 41%). H NMR (500 MHz, CD3OD): δ
7.62 (dt, J = 7.5, 3.5 Hz, 2H), 7.58−7.51 (m, 2H), 7.37 (t, J = 7.5 Hz,
1H), 7.25 (t, J = 7.5 Hz, 1H), 7.20 (d, J = 7.5 Hz, 1H), 7.13−7.07 (m,
3H), 6.93 (dt, J = 7.5, 2.5 Hz, 1H), 6.69 (dd, J = 7.5, 2.5 Hz, 1H),
6.60−6.54 (m, 2H), 5.05−5.00 (m, 1H), 4.15−4.00 (m, 2H). 13C
NMR (125 MHz, CD3OD): δ 161.37, 144.06, 131.34, 130.62, 129.88,
128.51, 128.38, 123.20, 122.77, 121.79, 116.23, 110.97, 110.09, 74.18,
73.46. LC-TOF (M + H+) calcd for C24H23N4O2S2 463.1257, found
463.1256.
N,N′-((Ethane-1,2-diylbis(oxy))bis(3,1-phenylene))bis-
(thiophene-2-carboximidamide) (11). To a solution of 3-nitro-
phenol (2 mmol) in DMF (20 mL) was added K2CO3 (2.2 mmol) and
1,3-dibromopropane (1 mmol). The suspension was then stirred for 6
h at 60 °C. The reaction mixture was then diluted with H2O (50 mL),
extracted with EtOAc, and washed with water and brine. The organic
layer was dried over Na2SO4 and concentrated in vacuo. The residue
was purified by column chromatography to yield 27 (yield 80%). 11
was synthesized by general procedures A and D using 27 as the
130.90, 130.43, 129.86, 129.79, 128.43, 128.37, 128.35, 125.79, 124.76,
123.65, 122.58, 58.73, 55.97, 33.01, 11.75. LC-TOF (M + H+) calcd
for C27H30N5S2 488.1937, found 488.1932.
N-(3-(((3-Fluorophenethyl)amino)methyl)phenyl)thiophene-
2-carboximidamide (16). 16 was synthesized by the same
procedures as those to prepare 13 using 3-fluorobenzaldehyde as the
1
starting material (yield 59%). H NMR (500 MHz, CD3OD): δ 8.01
1
(t, J = 4.0 Hz, 4H), 7.47 (t, J = 8.0 Hz, 2H), 7.33 (t, J = 4.0 Hz, 2H),
7.03 (dd, J = 8.0, 2.0 Hz, 2H), 6.97 (t, J = 2.0 Hz, 2H), 6.94 (dd, J =
8.0, 2.0 Hz, 2H), 4.43 (s, 4H). 13C NMR (125 MHz, CD3OD): δ
161.52, 157.72, 133.75, 133.12, 132.15, 129.64, 129.49, 118.15, 114.83,
112.12, 68.12. LC-TOF (M + H+) calcd for C24H23N4O2S2 463.1257,
found 463.1257.
N,N′-((Propane-1,3-diylbis(oxy))bis(3,1-phenylene))bis-
(thiophene-2-carboximidamide) (12). 12 was synthesized by the
same procedures as those to prepare 11 using 1,4-dibromobutane as
the starting material. 1H NMR (500 MHz, CD3OD): δ 8.09−7.99 (m,
4H), 7.49 (t, J = 8.0 Hz, 2H), 7.41−7.33 (m, 2H), 7.11−7.05 (dd, J =
8.0, 2.5 Hz, 2H), 7.03 (t, J = 2.5 Hz, 2H), 7.00 (dd, J = 8.0, 2.5 Hz,
starting material. H NMR (400 MHz, CD3OD): δ 8.18−8.04 (m,
2H), 7.81−7.69 (m, 2H), 7.65 (t, J = 7.5 Hz, 1H), 7.58−7.50 (m, 1H),
7.41−7.28 (m, 2H), 7.18−7.06 (m, 2H), 7.03−6.93 (m, 1H), 4.37 (s,
2H), 3.42−3.31 (m, 2H), 3.20−3.10 (m, 2H). 13C NMR (100 MHz,
CD3OD): δ 164.17, 161.74 (s), 139.31 (d, J = 7.5 Hz), 134.72, 134.45,
134.22, 133.50, 130.87, 130.55, 130.34 (d, J = 7.5 Hz), 128.66, 127.22,
126.47, 124.49, 124.46, 115.30 (d, J = 20 Hz), 113.56 (d, J = 20 Hz),
109.99, 50.38, 48.27, 31.49 (d, J = 2.0 Hz). LC-TOF (M + H+) calcd
for C20H21FN3S 354.1435, found 354.1441.
N-(3-((Ethyl(3-fluorophenethyl)amino)methyl)phenyl)-
thiophene-2-carboximidamide (17). 17 was synthesized by the
same procedures as those to prepare 15 using 3-fluorobenzaldehyde
K
dx.doi.org/10.1021/jm401252e | J. Med. Chem. XXXX, XXX, XXX−XXX