3
72 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
-Chloro-2-((3-ethyl-5-(2-(3-methylbenzo[d]thiazol-2(3H)-yl-
Yang et al.
þ
5
4.04 (s, 3H), 3.76 (s, 6H), 1.26 (t, J=7.0, Hz, 3H). MS (ESI )
m/z: 480.1 [M ].
þ
idene)ethylidene)-4-oxothiazolidin-2-ylidene)amino)-3-methyl-
benzo[d]thiazol-3-ium Chloride (7). Dark-blue solid, mp
2-((5-((3,5-Dimethylthiazol-2(3H)-ylideneamino)methylene)-
3-ethyl-4-oxothiazolidin-2-ylidene)methyl)-5-fluoro-3-methyl-
benzo[d]thiazol-3-ium Bromide (14). Dark-blue solid, mp
-
1
1
2
24-228 ꢀC. IR ν (KBr, cm ): 1519, 1456. H NMR (400
MHz, DMSO-d ) δ: 8.40 (br, 1H), 8.02-8.00 (m, 1H), 7.93 (d,
J=7.8 Hz, 1H), 7.88 (d, J=13.2 Hz, 1H), 7.84-7.81 (m, 1H),
.53 (t, J=7.2 Hz, 1H), 7.37 (t, J=8.0 Hz, 1H), 6.19 (d, J=13.2
Hz, 1H), 4.11 (q, J=7.2 Hz, 2H), 4.02 (s, 3H), 1.20 (t, J=7.2 Hz,
6
-
1
-1
251-253 ꢀC. UV-vis (MeOH): λ (nm) (log ε, L mol cm ):
-
1
7
526 (4.86), 362 (4.56). IR ν (neat, cm ): 2975, 1659, 1584, 1528,
1489, 1417, 1389, 1333, 1279, 1204, 1148, 1061, 993, 979, 939,
þ
þ
1
3
H). MS (ESI ) m/z: 499 [M ].
-((3-Ethyl-5-(2-(3-methylbenzo[d]thiazol-2(3H)-ylidene)ethyl-
idene)-4-oxothiazolidin-2-ylidene)methyl)-5-fluoro-3-methylbenzo-
893, 834, 755, 738. H NMR (400 MHz, DMSO-d ) δ: 8.33 (dd,
6
2
J = 8.9 and 5.2 Hz, 1H), 8.06 (s, 1H), 8.02 (dd, J = 9.9 and 2.4
Hz, 1H), 7.47 (dt, J = 8.9 and 2.4 Hz, 1H), 7.32 (d, J = 1.4 Hz,
1H), 6.74 (s, 1H), 4.15 (q, J = 7.0 Hz, 2H), 4.07 (s, 3H), 3.60 (s,
[
d]thiazol-3-ium Maleate (9). Dark-green solid, mp 260.1
-
1
-1
þ
-
260.8 ꢀC. UV-vis (H O): λ (nm) (log ε, L mol cm ): 526
3H), 2.30 (s, 3H), 1.24 (t, J = 7.0, Hz, 3H). MS (ESI ) m/z:
447.0 [M ].
2
-
1
þ
(
4.68), 355 (4.37). IR ν (neat, cm ): 2972, 2937, 2866, 1684,
1
9
526, 1472, 1376, 1360, 1346, 1314, 1275, 1198, 1055, 1032, 1013,
40, 891, 818, 747. H NMR (400 MHz, DMSO-d ) δ: 8.28 (dd,
6
2-((5-((3,4-Dimethylthiazol-2(3H)-ylideneamino)methylene)-
3-ethyl-4-oxothiazolidin-2-ylidene)methyl)-5-fluoro-3-methyl-
benzo[d]thiazol-3-ium Bromide (15). Dark-blue solid, mp
1
J=8.8 and 5.1 Hz, 1H), 7.99 (dd, J=9.9 and 1.9 Hz, 1H), 7.89 (d,
J=7.8 Hz, 1H), 7.75 (d, J=13.2 Hz, 1H), 7.59 (d, J=8.2 Hz, 1H),
-
1
-1
277-279 ꢀC. UV-vis (MeOH): λ (nm) (log ε, L mol cm ):
-
1
7
.49 (dd, J=17.0 and 8.3 Hz, 2H), 7.33 (t, J=7.6 Hz, 1H), 6.78
s, 1H), 6.03 (d, J=13.2 Hz, 3H), 4.19 (q, J=7.1 Hz, 2H), 4.06
526 (4.92), 362 (4.64). IR ν (neat, cm ): 2916, 1681, 1584, 1525,
1474, 1416, 1375, 1328,1276, 1199, 1157, 1062, 1033, 869, 838,
(
(
þ
1
749. H NMR (400 MHz, DMSO-d
s, 3H), 3.78 (s, 3H), 1.27 (t, J = 7.1 Hz, 3H). MS (ESI )
) δ: 8.34 (dd, J = 8.9 and
6
þ
m/z: 482.3 [M ].
-((3-Ethyl-5-(2-(3-methylbenzo[d]thiazol-2(3H)-ylidene)ethyl-
idene)-4-oxothiazolidin-2-ylidene)methyl)-5-fluoro-3-methyl-
benzo[d]thiazol-3-ium Methanesulfonate (10). Dark-green solid,
5.2 Hz, 1H), 8.18 (s, 1H), 8.03 (dd, J = 9.9 and 2.3 Hz, 1H), 7.48
(dt, J = 8.9 and 2.3 Hz, 1H), 6.83 (d, J = 1.1 Hz, 1H), 6.76 (s,
1H), 4.16 (q, J = 7.0 Hz, 2H), 4.08 (s, 3H), 3.62 (s, 3H), 2.30 (s,
2
þ
þ
3H), 1.25 (t, J = 7.0, Hz, 3H). MS (ESI ) m/z: 447.0 [M ].
2-((3-Ethyl-5-((1-methylpridin-2(1H)-ylideneamino)methyl-
ene)-4-oxothiazolidin-2-ylidene)methyl)-5-fluoro-3-methylbenzo-
[d]thiazol-3-ium Bromide (16). Dark-blue solid, mp 256-260 ꢀC.
-1
-1
mp >300 ꢀC. UV-vis (H
2
O): λ (nm) (log ε, L mol cm ): 524
-
1
(
4.83), 355 (4.52). IR ν (neat, cm ): 2971, 2937, 1682, 1525,
470, 1376, 1358, 1317, 1276, 1197, 1057, 1033, 939, 891, 819,
1
7
5
1
2
1
1
47. H NMR (400 MHz, DMSO-d
-1
-1
6
) δ: 8.29 (dd, J = 8.8 and
UV-vis (MeOH): λ (nm) (log ε, L mol cm ): 530 (4.74), 368
-1
.2 Hz, 1H), 8.01 (dd, J=9.9 and 2.0 Hz, 1H), 7.90 (d, J=7.8 Hz,
H), 7.76 (d, J=13.2 Hz, 1H), 7.61 (d, J=8.3 Hz, 1H), 7.49 (m,
H), 7.34 (t, J=7.8 Hz, 1H), 6.77 (s, 1H), 6.05 (d, J=13.2 Hz,
H), 4.19 (q, J=7.1 Hz, 2H), 4.05 (s, 3H), 3.77 (s, 3H), 2.29
(4.41). IR ν (neat, cm ): 2975, 1649, 1519, 1448, 1333, 1274,
1203, 1163, 1061, 994, 938, 898, 835, 796, 763. H NMR (400
1
MHz, DMSO-d ) δ: 8.7 (s, 1H), 8.29 (dd, J = 9.0 and 4.5 Hz,
6
2H), 7.99-7.88 (m, 2H), 7.65 (d, J = 9.0 Hz, 1H), 7.47-7.35
(m, 1H), 7.01 (t, J = 6.7, Hz, 1H), 6.68 (s, 1H), 4.17 (q, J = 7.0
Hz, 2H), 4.03 (s, 3H), 3.92 (s, 3H), 1.25 (t, J = 7.0, Hz, 3H). MS
þ
þ
(
s, 3H), 1.26 (t, J=7.1 Hz, 3H). MS (ESI ) m/z: 483.2 [M ].
-Fluoro-2-((3-(2-hydroxyethyl)-5-(2-(3-methylbenzo[d]thia-
zol-2(3H)-ylidene)ethylidene)-4-oxothiazolidin-2-ylidene)methyl)-
5
þ
þ
(ESI ) m/z: 427.1 [M ].
3
-methylbenzo[d]thiazol-3-ium Chloride (11). Dark-green solid,
Biology. In Vitro Anti-Leishmanial Assay. The Leishmania
donovani strain MHOM/ET/67/L82 was used. The strain was
maintained in the Syrian Golden hamster. Amastigotes were
collected from the spleen of an infected hamster and then grown
in axenic culture at 37 ꢀC in Cunningham SM medium, pH 5.4,
supplemented with 10% heat-inactivated fetal bovine serum
-
1
2
mp 277.0-277.7 ꢀC. UV-vis (H O): λ (nm) (log ε, L mol
cm ): 600 (3.95), 379 (3.50). IR ν (neat, cm ): 3668, 2921,
-1
-1
2
1
d
2
7
1
868, 1732, 1524, 1498, 1472, 1371, 1345, 1314, 1269, 1249, 1192,
1
145, 1059, 1033, 1015, 819, 751. H NMR (400 MHz, DMSO-
6
) δ: 8.29 (dd, J = 8.9 and 5.2 Hz, 1H), 7.98 (dd, J=10.0 and
(
FBS) under an atmosphere of 5% CO2 in air. Stock drug
solutions are prepared in 100% dimethyl sulfoxide (DMSO) at
0 mg/mL and heated or sonicated if necessary to dissolve the
.2 Hz, 1H), 7.89 (d, J=7.8 Hz, 1H), 7.74 (d, J=13.2 Hz, 1H),
.59 (d, J=8.2 Hz, 1H), 7.54-7.43 (m, 2H), 7.32 (t, J=7.7 Hz,
H), 6.93 (s, 1H), 6.02 (d, J=13.2 Hz, 1H), 5.19 (br, 1H), 4.78 (t,
1
sample. Assays were performed in 96-well flat-bottom micro-
titer plates, with each well containing 100 μL of culture medium
with 10 amastigotes from axenic culture with or without a serial
J = 4.4 Hz, 2H), 4.16 (q, J= 6.9, 6.9 Hz, 2H), 3.87 (t, J = 3.9
Hz, 2H), 3.78 (s, 3H), 1.24 (t, J = 7.1 Hz, 3H). MS (ESI )
m/z: 512.1 [M ].
þ
5
þ
drug dilution. After 72 h of incubation, the plates were inspected
under an inverted microscope to ensure growth of the controls
and sterile conditions. An amount of 10 μL of Alamar blue
(12.5 mg of resazurin dissolved in 100 mL of distilled water) was
then added to each well, and the plates were incubated for
another 2 h. The plates were then read with a microplate
fluorometer using an excitation wavelength of 536 nm and an
emission wavelength of 588 nm. The decrease of fluorescence
(i.e., inhibition) was expressed as a percentage of the fluore-
scence of control cultures and plotted against the drug concen-
trations. The IC50 value was calculated from the sigmoidal
inhibition curve by the software program.
2
-((Ethyl-5-(2-(4-methyloxazolo[4,5-b]pyridin-2(4H)-ylidene)-
ethylidene)-4-oxothiazolidin-2-ylidene)methyl)-5-fluoro-3-methyl-
benzo[d]thiazol-3-ium Bromide (12). Dark-green solid, mp
86-288 ꢀC. UV-vis (MeOH): λ (nm) (log ε, L mol cm ):
16 (5.00), 382 (4.33). IR ν (neat, cm- ): 2982, 2819, 1771, 1747,
715, 1697, 1682, 1648, 1542, 1524, 1489, 1338, 1227, 1033, 835.
-1
-1
2
6
1
1
1
6
H NMR (400 MHz, DMSO-d ) δ: 8.21 (dd, J = 8.5 and 5.2
Hz, 1H), 8.18-8.16 (m, 1H), 8.13 (d, J = 5.2 Hz, 1H),
8
7
(
.05-7.79 (m, 2H), 7.37 (dt, J = 8.7, 8.5, and 1.9 Hz, 1H),
.27-7.10 (m, 1H), 6.67 (s, 1H), 5.47 (d, J = 13.6 Hz, 1H), 4.15
q, J = 7.2 Hz, 2H), 4.08 (s, 3H), 3.99 (s, 3H), 1.24 (t, J = 7.2,
þ
þ
Hz, 3H). MS (ESI ) m/z: 467.1 [M ].
-((5-((1,3-Dimethyl-1H-benzo[d]imidazol-2(3H)-ylideneamino)-
methylene)-3-ethyl-4-oxothiazolidin-2-ylidene)methyl)-5-fluoro-
2
Cytotoxicity Assay. The L-6 rat skeletal myoblast cell line was
grown in RPMI 1640 medium supplemented with 1% L-gluta-
mine (200 mM) and 10% FBS in T-25 tissue culture flasks at
3
-methylbenzo[d]thiazol-3-ium Bromide (13). Dark-blue solid,
-
1
mp 274-276 ꢀC. UV-vis (MeOH): λ (nm) (log ε, L mol
37 ꢀC in 5% CO in air. The cultures were subpassaged three
2
-
1
-1
cm ): 512 (4.76), 350 (4.57). IR ν (neat, cm ): 1680,
times a week, using trypsin to detach the cells, and split in a 1:2
or 1:3 ratio, depending on the density of the parent culture.
Samples were cryopreserved at a low passage number. Stock
drug solutions were prepared in 100% DMSO at 10 mg/mL and
heated or sonicated if necessary to dissolve the sample. Assays
were performed in 96-well microtiter plates, each well receiving
1
1
658, 1515, 1473, 1425, 1350, 1331, 1277, 1201, 1184, 1155,
134, 1061, 997, 940, 884, 831, 756. H NMR (400 MHz,
DMSO-d ) δ: 8.75 (s, 1H), 8.30 (dd, J=8.9 and 5.5 Hz, 1H),
.97 (dd, J=10.0 and 2.5 Hz, 1H), 7.63 (dd, J=5.5 and 2.5 Hz,
H), 7.46-7.39 (m, 3H), 6.71 (s, 1H), 4.18 (q, J=7.0 Hz, 2H),
1
6
7
2