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S. LASKAR ET AL.
lymphocytes, unlike cisplatin which induced necrotic cell death 72 h, implying a greater degree of antiproliferative selectivity
towards malignant cell lines than with lymphocytes. In silico ADME
profiling study revealed that compound 9e has the potential to be
developed as oral drug candidate (Table 4).
(Figure 4). In the preliminary apoptosis experiment CaSki, MDA-
MB-231 and SK-Lu-1 cultures were stimulated at the level of their
determined IC50 values and the morphological changes, chromatin
condensation including the formation of apoptotic bodies were
determined through staining with fluorochrome 40,6-diamidino-2-
phenylindole (DAPI). Compact nuclei and apoptotic bodies were
clearly observed in the cultures (Figure 5). The condensed chroma-
tin in treated cells suggested that compound 9e induced cell
death by apoptosis in the concerned cancer cell lines. In the pre-
sent study, we had detected active caspase-3 by immunodetec-
tion. Figure 5 shows that compound 9e induced the expression of
active caspase-3 in CaSki, MDA-MB-231 and SK-Lu-1 cultures,
implying that apoptosis could be triggered through a caspase
dependent process. It is well known that during chemotherapy
the immune system is usually affected. Thus to evaluate the
selectivity, the proliferation of enriched lymphocyte population
(ELP) was evaluated with compound 9e using CFSE-labelling assay
(Figure 6). The results indicated that with compound 9e, prolifera-
tive potential of lymphoid cells was not negatively affected after
Table 3. Antiproliferative activities of the synthesized (1S,4S)-N-Boc-2,5-diazabi-
cyclo[2.2.1]heptane-dithiocarbamates (9a–9g).
IC50 (mg mLꢂ1
)
Compound
CaSki
MDA-MB231
SK-Lu-1
9a
9b
9c
9d
346
294
305
214
28
150
69
100
100
18
300
57
120
87
9e
20
9f
348
137
1.67
227
50
2.37
237
40
1.36
9g
Cisplatina
All the experimental results are the average of three independent experiments.
aThe assay was performed with a commercially available sample of Cisplatin,
purchased from Sigma-Aldrich.
Table 4. In silico prediction of physicochemical pharmacokinetic properties24
.
Table 1. Experimental data of the compounds 9a–9g.
Code
miLogPa
nONb
nOHNHc
n-Violationsd
Nrotbe
MWf
SL.
no
Time
(h)
Isolated
yield (%) Melting point
Rule
9e
ꢃ5
<10
4
<5
0
ꢃ1
–
6
<500
342.47
20
Product
Optical rotation, [a]D
ꢂ193.9 (c 1, CH2Cl2)
4.55
0
1
2
3
4
5
6
7
9a
9b
9c
9d
9e
9f
10
10
11
12
10
10
10
70
68
60
55
63
75
71
109–110 ꢀC
Viscous liquid
amiLogP: logarithm of partition coefficient of compound between n-octanol and
water.
ꢂ150.8 (c 1.025, CH2Cl2)
Viscous liquid ꢂ166.84 (c 1.04, CH2Cl2)
bn-ON acceptors: number of hydrogen bond acceptors.
cn-OHNH donors: number of hydrogen bonds donors.
dn-violations: number of violations according to Lipinski’s rule.
eNrotb: number of rotatable bonds.
228–230 ꢀC
Viscous liquid
98–103 ꢀC
ꢂ79.61 (c 0.336, DMSO)
ꢂ140.4 (c 1.04, CH2Cl2)
ꢂ178.3 (c 178.3, CH2Cl2)
ꢂ122.7 (c 1.046, CH2Cl2)
9g
Viscous liquid
fMW: molecular weight.
Table 2. Spectral data of the compounds 9a–9g.
7
1H-NMR (300 MHz, CDCl3, ppm): d ¼ 4.33 (0.5H, s, –CH), 4.21 (0.5H, s, –CH), 3.60 (1H, s, –CH), 3.28–3.22 (1H, m, –CH2), 3.11-3.05 (1H, m, –CH2), 2.97–2.87
(2H, m, –CH2), 2.14 (1H, s, –NH), 1.62-1.56 (2H, m, –CH2), 1.35 (9H, brs, –CH3); 13C-NMR (75 MHz, CDCl3, ppm): d ¼ 154.43, 154.16, 79.39, 79.29, 57.16,
56.51, 56.11, 55.98, 52.57, 52.44, 37.03, 36.80, 28.5120
9a
1H-NMR (400 MHz, DMSO-d6, ppm): d ¼ 5.42 (0.5H, s, –CH), 4.99 (0.5H, s, –CH), 4.55–4.29 (1H, m, –CH), 3.74–3.63 (1H, m, –CH2), 3.49–3.40 (1H, m, –CH2),
3.28–3.13 (2H, m, –CH2), 2.58–2.56 (3H, m, –CH3), 2.02–1.75 (2H, m, –CH2), 1.40–1.38 (9H, m, –CH3); 13C-NMR (100 MHz, DMSO-d6, ppm): d ¼ 193.36
(C ¼ S), 153.44 (C ¼ O), 64.30, 63.74, 62.52, 62.16, 61.96, 61.40, 58.58, 58.26, 57.33, 57.16, 56.62, 56.36, 56.21, 56.00, 55.70, 54.99, 54.04, 53.63, 53.31,
52.37, 52.10, 51.76, 37.48, 36.94, 36.25, 35.74, 28.11, 28.06, 18.69, 18.56; MS (FABþ): m/z calculated for C12H20N2O2S2 [M þ H]þ: 289, found: 289
9b
9c
9d
9e
1H-NMR (400 MHz, CDCl3, ppm): d ¼ 7.31–7.16 (5H, m, –ArH), 5.59 (0.5H, s, –CH), 4.89 (0.5H, s, –CH), 4.53–4.43 (3H, m, –CH, –CH2), 3.89–3.31 (4H, m, –CH2),
1.90–1.84 (2H, m, –CH2), 1.39–1.37 (9H, m, –CH3); 13C-NMR (100 MHz, CDCl3, ppm): d ¼ 193.78 (C ¼ S), 192.78 (C ¼ S), 154.06 (C ¼ O), 136.14, 129.35,
128.69, 127.62, 80.32, 64.54, 63.89, 62.38, 62.28, 62.18, 61.59, 58.35, 57.55, 56.61, 56.21, 55.23, 52.34, 51.98, 41.25, 41.12, 38.02, 37.55, 36.91, 36.49, 28.50;
MS (FABþ): m/z calculated for C18H24N2O2S2 [M þ H]þ: 365, found: 365
1H-NMR (400 MHz, CDCl3, ppm): d ¼ 8.51 (1H, d, J ¼ 4 Hz, –ArH), 7.60 (1H, td, J ¼ 7.6 Hz, 2 Hz, –ArH), 7.45 (1H, m, –ArH), 7.15–7.12 (1H, m, –ArH), 5.62 (0.5H,
s, –CH), 5.01 (0.5H, s, –CH), 4.80–4.47 (3H, m, –CH, –CH2), 3.89–3.37 (4H, m, –CH2), 1.95–1.89 (2H, m, –CH2), 1.42–1.40 (9H, m, –CH3); 13C-NMR (100 MHz,
CDCl3, ppm): d ¼ 193.56 (C ¼ S), 156.88, 154.11 (C ¼ O), 149.65, 136.83, 123.97, 122.47, 80.44, 64.84, 64.23, 62.62, 62.56, 62.35, 61.78, 58.50, 57.64, 56.70,
56.29, 55.31, 52.41, 52.09, 52.01, 42.63, 38.11, 37.67, 37.01, 36.59, 28.58; MS (FABþ): m/z calculated for C17H23N3O2S2 [M þ H]þ: 366, found: 366
1H-NMR (400 MHz, CDCl3, ppm): d ¼ 7.52 (2H, brs, –ArH), 7.21–7.19 (2H, m, –ArH), 5.62 (0.5H, s, –CH), 4.95 (0.5H, s, –CH), 4.91–4.50 (3H, m, –CH, –CH2),
3.95–3.31 (4H, m, –CH2), 1.95–1.85 (2H, m, –CH2), 1.44–1.40 (9H, m, –CH3); 13C-NMR (100 MHz, CDCl3, ppm): d ¼ 193.61 (C ¼ S), 154.00 (C ¼ O), 151.47,
122.89, 115.56, 80.66, 65.60, 65.00, 63.18, 62.97, 62.37, 58.95, 57.56, 56.67, 56.21, 55.23, 52.32, 51.93, 38.13, 37.69, 37.07, 36.63, 33.30, 33.14, 28.56; MS
(FABþ): m/z calculated for C19H24N4O2S2 [M þ H]þ: 405, found: 405
1H-NMR (400 MHz, CDCl3, ppm): d ¼ 7.31 (1H, s, –ArH), 7.22-7.15 (3H, m, –ArH), 5.59 (0.5H, s, –CH), 4.90 (0.5H, s, –CH), 4.57–4.40 (3H, m, –CH, –CH2),
3.89–3.72 (1H, m, –CH2), 3.63–3.33 (3H, m, –CH2), 1.92–1.85 (2H, m, –CH2), 1.40–1.38 (9H, m, –CH3); 13C-NMR (100 MHz, CDCl3, ppm): d ¼ 193.26 (C ¼ S),
192.19 (C ¼ S), 154.13 (C ¼ O), 138.65, 138.60, 134.43, 129.95, 129.41, 127.84, 127.82, 127.58, 80.46, 64.81, 64.16, 62.60, 62.51, 62.32, 61.73, 58.45, 56.66,
56.26, 55.29, 52.41, 52.05, 40.43, 40.29, 38.11, 37.65, 36.98, 36.57, 28.58; MS (FABþ) m/z (%) ¼ 399 [Mþ], 343, 341, 273, 241, 140, 185, 141, 125, 57; MS
(FABþ): m/z calculated for C18H23ClN2O2S2 [M þ H]þ: 399, found: 399
9f
1H-NMR (400 MHz, DMSO-d6, ppm): d ¼ 7.45–7.41 (2H, m, –ArH), 7.17–7.10 (2H, m, –ArH), 5.43 (0.5H, s, –CH), 5.00 (0.5H, s, –CH), 4.60–4.29 (3H, m, –CH,
–CH2), 3.78–3.65 (1H, m, –CH2), 3.48–3.37 (1H, m, –CH2), 3.34–3.12 (2H, m, –CH2), 2.00–1.75 (2H, m, –CH2), 1.40–1.38 (9H, m, –CH3); 13C-NMR (100 MHz,
DMSO-d6, ppm): d ¼ 191.66 (C ¼ S), 162.59, 160.17, 153.36 (C ¼ O), 132.82, 131.09, 131.01, 115.31, 115.10, 79.25, 79.13, 64.45, 63.88, 62.60, 62.25, 62.12,
61.54, 58.58, 58.31, 57.27, 56.31, 55.92, 54.92, 52.38, 52.05, 51.66, 38.86, 37.42, 36.90, 36.16, 35.65, 28.07, 28.01; MS (FABþ): m/z calculated for
C18H23FN2O2S2 [M þ H]þ: 383, found: 383
9g
1H-NMR (400 MHz, CDCl3, ppm): d ¼ 7.45–7.24 (5H, m, –ArH), 5.65–5.61 (0.3H, m), 5.39 (0.24H, d, J ¼ 8 Hz), 5.02–4.97 (0.8H, m), 4.70–4.35 (1H, m), 4.15–3.27
(7H, m), 1.98–1.73 (2H, m, –CH2), 1.43–1.38 (9H, m, –CH3); 13C-NMR (100 MHz, CDCl3, ppm): d ¼ 193.18 (C ¼ S), 154.10 (C ¼ O), 143.03, 129.06, 128.97,
128.74, 128.66, 128.27, 128.15, 128.01, 127.98, 126.00, 125.97, 80.54, 73.48, 66.52, 66.27, 65.99, 65.04, 64.96, 64.84, 64.39, 62.70, 62.54, 61.95, 58.78,
58.67, 58.55, 57.60, 57.12, 56.97, 56.66, 56.30, 55.33, 55.26, 54.17, 53.74, 53.58, 52.35, 52.09, 51.99, 45.12, 44.93, 44.77, 44.63, 38.09, 37.64, 37.02, 36.62,
28.60; MS (FABþ): m/z calculated for C19H26N2O3S2 [M þ H]þ: 395, found: 395