Abstract
In order to study the structure-activity relationships of xanthene derivatives, four series of N-substituted 14-aryl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide derivatives were synthesized. The structures of all compounds were identified by 1H-NMR, HR-MS and IR spectra, in which compounds 6a–h were further identified by 13C-NMR spectra. The in vitro antitumor activity of the synthesized compounds was tested by MTT assay. Most of them displayed strong inhibitory activity on human hepatocellular carcinoma cell lines (SK-HEP-1, HepG2 and SMMC-7721 cells) and acute promyelocytic leukemia NB4 cells. Compounds 6c–6e exhibited significant inhibitory activity against NB4 cells with IC50 values of 0.52 μM and 0.76 μM, respectively, much lower than 5.31 μM of the positive control As2O3.
Keywords: synthesis design; cytotoxicity; dibenzo[a,j]xanthenes; NMR spectroscopy
1. Introduction
The benzoxanthenes have attracted considerable interest due to their biological and pharmacological activities such as antiviral [1], antibacterial [2], anti-inflammation [3], antitumor [4] and other usages in photodynamic therapy [5], and the antagonizing paralysis induced by zoxazolamine [6].
In view of the great importance of benzoxanthenes, the preparation of which has been a hot research topic, there have been many research reports in recent years [7,8,9,10,11]. So far the synthesis of dibenzo[a,j]xanthene has been mostly focused on the modification of the 14-position of the molecule, with other positions rarely reported in the literature, and especially the 3 and the 11 positions. In our previous work, we synthesized two series of dibenzo[a,j]xanthene-3,11-substituted compounds bearing a 2-hydroxyethyl group on the nitrogen atom (1a–i and 2a–c, Figure 1) [12]. The results of in vitro antitumor activity experiments revealed that compounds 1a–i and 2a–c exhibit remarkable inhibitory activity toward a wide range of human tumor cell lines. Furthermore, the amide derivatives 1a–c exhibit a stronger inhibitory effect than the amine derivatives 2a–c on tumor cell lines, implying that the amide group of dibenzo[a,j]xanthene derivatives is more critical than the amine group for improving the inhibitory activity toward tumor cells.
Figure 1.
Structures of compounds 1a–i and 2a–c.
These results prompted us to synthesize new analogues of compounds 1a–i containing different substituted amide groups, in order to clarify the structure-activity relationships (SARs) of xanthene analogues. Herein, we first describe the synthesis of derivatives 5a–h, 6a–h, 7a–h and 8a–h (Figure 2), and then report the screening results on their cytotoxicity against four cancer cell lines.
Figure 2.
Structures of compounds 5a–9d.
2. Results
2.1. Chemistry
The synthetic route for the carboxamide derivatives 5a–h, 6a–h, 7a–h and 8a–h is outlined in Scheme 1.
Scheme 1.
Synthesis of compounds 5a–h, 6a–h, 7a–h and 8a–h. Reagents and conditions: (I) See References [12,13,14]; (II) 1. Compounds 5a–5h: NH3, CHCl3, r.t., 2–3 h; 2. Compounds 6a–6h: CH3NH2, CHCl3, r.t., 2–3 h; 3. Compounds 7a–7h: CH3CH2CH2NH2, CHCl3, 2–3 h, r.t.; 4. Compounds 8a–8h: (CH3)2CHCH2NH2, CHCl3, r.t., 2–3 h.
14-Phenyl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5a): White solid. Yield 84.2%. m.p. 229–231 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.74 (d, J = 8.9 Hz, 2H, H-1, 13), 8.49 (s, 2H, H-4, 10), 8.10 (s, 2H, CONH × 2), 8.04 (overlapping t, J = 8.9 Hz, 4H), 7.72–7.57 (m, 4H), 7.44 (s, 2H, CONH × 2), 7.15 (t, J = 7.6 Hz, 2H, H-3′, 5′), 6.98 (t, J = 7.2 Hz, 1H, H-4′), 6.79 (s, 1H, H-14). IR (KBr) ν: 3422, 1655, 1623, 1595, 1466, 1396, 1244, 1081 cm−1. HR-MS (ESI) calcd for C29H21N2O3 [M + H]+ 445.1552, found 445.1559.
14-(2-Fluorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5b): White solid. Yield 83.7%. m.p. 229–230 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.51 (d, J = 1.5 Hz, 2H, H-4, 10), 8.47 (d, J = 9.0 Hz, 2H, H-1, 13), 8.10 (s, 2H, CONH × 2), 8.07(dd, J = 9.0, 1.7 Hz, 2H, H-2, 12), 8.06 (d, J = 9.0 Hz, 2H, H-5, 9), 7.63 (d, J = 8.9 Hz, 2H, H-6, 8), 7.62–7.57 (m, 1H, H-6′), 7.45 (s, 2H, CONH × 2), 7.14–6.97 (m, 3H, H-3′, 4′, 5′), 6.90 (s, 1H, H-14). IR (KBr) ν: 3409, 1658, 1624, 1595, 1467, 1397, 1256, 1245 cm−1. HR-MS (ESI) calcd for C29H20FN2O3 [M + H]+ 463.1458, found 463.1455.
14-(4-Fluorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5c): White solid. Yield 87.9%. m.p. 241–243 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.74 (d, J = 9.0 Hz, 2H, H-1, 13), 8.51 (d, J = 1.6 Hz, 2H, H-4, 10), 8.12 (s, 2H, CONH × 2), 8.06 (dd, J = 9.0, 1.7 Hz, 2H, H-2, 12), 8.04 (d, J = 8.9 Hz, 2H, H-5, 9), 7.72–7.62 (m, 2H, H-2′, 6′), 7.63 (d, J = 8.9 Hz, 2H, H-6, 8), 7.45 (s, 2H, CONH × 2), 6.99 (t, J = 8.9 Hz, 2H, H-3′, 5′), 6.82 (s, 1H, H-14). IR (KBr) ν: 3394, 3157, 3049, 1655, 1623, 1506, 1466, 1401, 1245 cm−1. HR-MS (ESI) calcd for C29H20FN2O3 [M + H]+ 463.1458, found 463.1469.
14-(2-Chlorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5d): White solid. Yield 82.5%. m.p. 235–236 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.65 (d, J = 9.0 Hz, 2H, H-1, 13), 8.51 (d, J = 1.5 Hz, 2H, H-4, 10), 8.10 (s, 2H, CONH × 2), 8.08 (dd, J = 9.0, 1.7 Hz, 2H, H-2, 12), 8.07 (d, J = 9.0 Hz, 2H, H-5, 9), 7.64 (d, J = 8.9 Hz, 2H, H-6, 8), 7.55 (d, J = 7.4 Hz, 1H, H-6′), 7.45 (s, 2H, CONH × 2), 7.34 (dd, J = 8.0, 1.1 Hz, 1H, H-3′), 7.15 (t, J = 7.0 Hz, 1H, H-5′), 7.07 (td, J = 7.8, 1.6 Hz, 1H, H-4′), 6.88 (s, 1H, H-14). IR (KBr) ν: 3377, 3192, 1657, 1623, 1594, 1467, 1396, 1254 cm−1. HR-MS (ESI) calcd for C29H20ClN2O3 [M + H]+ 479.1162, found 479.1170.
14-(4-Chlorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5e): White solid. Yield 82.3%. m.p. 231–233 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.72 (d, J = 8.9 Hz, 2H, H-1, 13), 8.50 (d, J = 1.6 Hz, 2H, H-4, 10), 8.11 (s, 2H, CONH × 2), 8.06 (dd, J = 8.9, 1.6 Hz, 2H, H-2, 12), 8.05 (d, J = 8.9 Hz, 2H, H-5, 9), 7.65 (d, J = 8.6 Hz, 2H, H-2′, 6′), 7.64 (d, J = 8.9 Hz, 2H, H-6, 8), 7.45 (s, 2H, CONH × 2), 7.23 (d, J = 8.6 Hz, 2H, H-3′, 5′), 6.82 (s, 1H, H-14). IR (KBr) ν: 3396, 3185, 1656, 1623, 1593, 1466, 1398, 1253 cm−1. HR-MS (ESI) calcd for C29H20ClN2O3 [M + H]+ 479.1162, found 479.1168.
14-(3-Nitrophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5f): White solid. Yield 81.8%. m.p. 225–227 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.81 (d, J = 9.0 Hz, 2H, H-1, 13), 8.56 (t, J = 1.9 Hz, 1H, H-2′), 8.51 (d, J = 1.6 Hz, 2H, H-4, 10), 8.12 (s, 2H, CONH × 2), 8.11–8.06 (m, 5H), 7.87 (br.d, J = 7.6 Hz, 1H, H-4′), 7.68 (d, J = 8.9 Hz, 2H, H-6, 8), 7.48 (t, J = 8.1 Hz, 1H, H-5′), 7.46 (s, 2H, CONH × 2), 7.04 (s, 1H, H-14). IR (KBr) ν: 3434, 3186, 1664, 1622, 1594, 1521, 1465, 1397, 1348, 1253 cm−1. HR-MS (ESI) calcd for C29H20N3O5 [M + H]+ 490.1403, found 490.1411.
14-(4-Nitrophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5g): White solid. Yield 83.7%. m.p. 238–240 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.75 (d, J = 9.0 Hz, 2H, H-1, 13), 8.51 (d, J = 1.5 Hz, 2H, H-4, 10), 8.12 (s, 2H, CONH × 2), 8.10-8.02 (m, 6H), 7.93 (d, J = 8.9 Hz, 2H, H-2′, 6′), 7.67 (d, J = 8.9 Hz, 2H, H-6, 8), 7.46 (s, 2H, CONH × 2), 7.00 (s, 1H, H-14). IR (KBr) ν: 3382, 3191, 1661, 1623, 1607, 1594, 1516, 1466, 1397, 1345, 1254 cm−1. HR-MS (ESI) calcd for C29H20N3O5 [M + H]+ 490.1403, found 490.1398.
14-(4-Methylphenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5h): White solid. Yield 84.2%. m.p. 229–231 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.72 (d, J = 9.0 Hz, 2H, H-1, 13), 8.50 (d, J = 1.4 Hz, 2H, H-4, 10), 8.12 (s, 2H, CONH × 2), 8.05 (dd, J = 9.0, 1.6 Hz, 2H, H-2, 12), 8.02 (d, J = 9.0 Hz, 2H, H-5, 9), 7.62 (d, J = 8.9 Hz, 2H, H-6, 8), 7.50 (d, J = 8.1 Hz, 2H, H-2′, 6′), 7.45 (s, 2H, CONH × 2), 6.95 (d, J = 8.0 Hz, 2H, H-3′, 5′), 6.74 (s, 1H, H-14), 2.05 (s, 3H, CH3). IR (KBr) ν: 3398, 3193, 1656, 1623, 1594, 1466, 1398, 1255, 1244 cm−1. HR-MS (ESI) calcd for C30H23N2O3 [M + H]+ 459.1709, found 459.1701.
N3,N11-Dimethyl-14-phenyl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6a): White solid. Yield 87.1%. m.p. >300 °C. 1H-NMR (300 MHz, DMSO-d6) δ (in ppm): 8.74 (d, J = 9.0 Hz, 2H, H-1, 13), 8.63 (q, J = 4.5 Hz, 2H, CONH × 2), 8.45 (br.s, 2H, H-4, 10), 8.03 (overlapping d, J = 8.9 Hz, 4H), 7.62 (overlapping d, J = 8.7 Hz, 4H), 7.13 (t, J = 7.6 Hz, 2H, H-3′, 5′), 6.96 (t, J = 7.3 Hz, 1H, H-4′), 6.77 (s, 1H, H-14), 2.82 (d, J = 4.4 Hz, 6H, CH3 × 2). 13C-NMR (75 MHz, DMSO-d6) δ (in ppm): 166.9, 149.4, 145.8, 132.6, 131.0, 130.6, 130.4, 129.0, 128.6, 128.4, 126.9, 125.4, 124.1, 118.9, 118.0, 36.9, 26.8. IR (KBr) ν: 3434, 1638, 1620, 1546, 1464, 1400, 1251 cm−1. HR-MS (ESI) calcd for C31H25N2O3 [M + H]+ 473.1865, found 473.1869.
N3,N11-Dimethyl-14-(2-fluorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6b): White solid. Yield 85.4%. m.p. >300 °C. 1H-NMR (300 MHz, DMSO-d6) δ (in ppm): 8.61 (br.q, J = 4.3 Hz, 2H, CONH × 2), 8.44 (overlapping d, J = 5.5 Hz, 4H), 8.03 (overlapping t, J = 7.5 Hz, 4H), 7.58 (overlapping t, J = 8.6 Hz, 3H), 7.13–6.92 (m, 3H), 6.86 (s, 1H, H-14), 2.82 (d, J = 4.1 Hz, 6H, CH3 × 2). 13C-NMR (75 MHz, DMSO-d6) δ (in ppm): 166.8, 159.0 (d, J = 244.3 Hz), 149.6, 132.6, 131.9 (d, J = 13.3 Hz), 131.4 (d, J = 3.2 Hz), 131.0, 131.0, 130.3, 129.5 (d, J = 8.3 Hz), 128.8, 125.7, 125.7, 123.0, 118.9, 116.3 (d, J = 22.5 Hz), 115.7, 31.6, 26.8. IR (KBr) ν: 3358, 1641, 1622, 1542, 1488, 1465, 1400, 1310, 1253 cm−1. HR-MS (ESI) calcd for C31H24FN2O3 [M + H]+ 491.1771, found 491.1778.
N3,N11-Dimethyl-14-(4-fluorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6c): White solid. Yield 86.6%. m.p. >300 °C. 1H-NMR (300 MHz, DMSO-d6) δ (in ppm): 8.74 (d, J = 8.9 Hz, 2H, H-1, 13), 8.68–8.57 (m, 2H, CONH × 2), 8.45 (br.s, 2H, H-4, 10), 8.03 (overlapping d, J = 8.9 Hz, 4H), 7.70–7.58 (m, 4H), 6.97 (t, J = 8.7 Hz, 2H, H-3′, 5′), 6.81 (s, 1H, H-14), 2.82 (d, J = 4.3 Hz, 6H, CH3 × 2). 13C-NMR (75 MHz, DMSO-d6) δ (in ppm): 166.9, 161.0 (d, J = 243.3 Hz), 149.3, 142.0, 132.6, 131.1, 130.8, 130.5, 130.2 (d, J = 8.0 Hz), 128.7, 125.5, 124.0, 118.9, 117.8, 115.7 (d, J = 21.2), 36.0, 26.8. IR (KBr) ν: 3306, 1642, 1549, 1506, 1464, 1401, 1252 cm−1. HR-MS (ESI) calcd for C31H24FN2O3 [M + H]+ 491.1771, found 491.1762.
N3,N11-Dimethyl-14-(2-chlorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6d): White solid. Yield 89.3%. m.p. >300 °C. 1H-NMR (300 MHz, DMSO-d6) δ (in ppm): 8.60 (overlapping d, J = 8.7 Hz, 4H), 8.45 (br.s, 2H, H-4, 10), 8.04 (overlapping d, J = 8.8 Hz, 4H), 7.59 (d, J = 8.9 Hz, 2H, H-6, 8), 7.49 (d, J = 7.3 Hz, 1H, H-6′), 7.32 (d, J = 7.7 Hz, 1H, H-3′), 7.11 (t, J = 7.4 Hz, 1H, H-5′), 7.03 (t, J = 7.2 Hz, 1H, H-4′), 6.79 (s, 1H, H-14), 2.82 (d, J = 4.2 Hz, 6H, CH3 × 2). 13C-NMR (75 MHz, DMSO-d6) δ (in ppm): 166.8, 149.7, 142.8, 132.7, 132.3, 131.1, 131.0, 130.6, 130.5, 130.4, 129.2, 128.8, 128.7, 125.5, 123.6, 119.0, 116.7, 35.2, 26.8. IR (KBr) ν: 3428, 1640, 1550, 1465, 1400, 1252 cm−1. HR-MS (ESI) calcd for C31H24ClN2O3 [M + H]+ 507.1475, found 507.1481.
N3,N11-Dimethyl-14-(4-chlorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6e): White solid. Yield 82.7%. m.p. >300 °C. 1H-NMR (300 MHz, DMSO-d6) δ (in ppm): 8.71 (d, J = 9.0 Hz, 2H, H-1, 13), 8.56 (br.q, J = 4.6 Hz, 2H, CONH × 2), 8.44 (br.s, 2H, H-4, 10), 8.03 (overlapping t, J = 8.4 Hz, 4H), 7.67–7.58 (m, 4H), 7.21 (d, J = 8.5 Hz, 2H, H-3′, 5′), 6.81 (s, 1H, H-14), 2.83 (d, J = 4.5 Hz, 6H, CONCH3 × 2). 13C-NMR (75 MHz, DMSO-d6) δ (in ppm): 166.9, 149.4, 144.7, 132.5, 131.6, 131.2, 130.8, 130.5, 130.1, 129.0, 128.6, 125.5, 124.0, 118.9, 117.5, 36.2, 26.8. IR (KBr) ν: 3350, 1642, 1548, 1488, 1465, 1400, 1253 cm−1. HR-MS (ESI) calcd for C31H24ClN2O3 [M + H]+ 507.1475, found 507.1483.
N3,N11-Dimethyl-14-(3-nitrophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6f): White solid. Yield 84.9%. m.p. >300 °C. 1H-NMR (300 MHz, DMSO-d6) δ (in ppm): 8.80 (d, J = 9.0 Hz, 2H, H-1, 13), 8.64–8.55 (m, 3H), 8.46 (s, 2H, H-4, 10), 8.10–8.00 (m, 5H), 7.86 (br.d, J = 8.0 Hz, 1H, H-4′), 7.67 (d, J = 8.9 Hz, 2H, H-6, 8), 7.46 (t, J = 8.0 Hz, 1H, H-5′), 7.02 (s, 1H, H-14), 2.82 (d, J = 4.4 Hz, 6H, CONCH3 × 2). 13C-NMR (75 MHz, DMSO-d6) δ (in ppm): 166.8, 149.5, 148.3, 147.7, 134.8, 132.5, 131.2, 131.2, 130.7, 130.5, 128.8, 125.7, 123.8, 122.5, 122.2, 119.0, 117.0, 36.3, 26.8. IR (KBr) ν: 3449, 3352, 1641, 1546, 1524, 1464, 1401, 1350, 1348, 1255 cm−1. HR-MS (ESI) calcd for C31H24N3O5 [M + H]+ 518.1716, found 518.1722.
N3,N11-Dimethyl-14-(4-nitrophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6g): White solid. Yield 84.2%. m.p. 285–287 °C. 1H-NMR (300 MHz, DMSO-d6) δ (in ppm): 8.75 (d, J = 8.9 Hz, 2H, H-1, 13), 8.60 (br.d, J = 4.3 Hz, 2H, CONH × 2), 8.45 (s, 2H, H-4, 10), 8.12–7.98 (m, 6H), 7.92 (d, J = 8.6 Hz, 2H, H-2′, 6′), 7.65 (d, J = 8.9 Hz, 2H, H-6, 8), 6.99 (s, 1H, H-14), 2.82 (d, J = 4.0 Hz, 6H, CH3 × 2). 13C-NMR (75 MHz, DMSO-d6) δ (in ppm): 166.8, 152.8, 149.4, 146.4, 132.5, 131.2, 131.2, 130.5, 129.5, 128.7, 125.7, 124.3, 123.9, 119.0, 116.7, 36.7, 26.8. IR (KBr) ν: 3353, 1645, 1550, 1523, 1465, 1399, 1344, 1255 cm−1. HR-MS (ESI) calcd for C31H24N3O5 [M + H]+ 518.1716, found 518.1710.
N3,N11-Dimethyl-14-(4-methylphenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6h): White solid. Yield 85.7%. m.p. >300 °C.1H-NMR (300 MHz, DMSO-d6) δ (in ppm): 8.71 (d, J = 9.0 Hz, 2H, H-1, 13), 8.58 (q, J = 4.5 Hz, 2H, CONH × 2), 8.43 (d, J = 1.4 Hz, 2H, H-4, 10), 8.04–7.96 (m, 4H), 7.60 (d, J = 8.9 Hz, 2H, H-6, 8), 7.48 (d, J = 8.1 Hz, 2H, H-2′, 6′), 6.92 (d, J = 8.0 Hz, 2H, H-3′, 5′), 6.72 (s, 1H, H-14), 2.82 (d, J = 4.5 Hz, 6H, CONCH3 × 2), 2.03 (s, 3H, Ar-CH3). 13C-NMR (75 MHz, DMSO-d6) δ (in ppm): 166.9, 149.3, 142.9, 136.0, 132.6, 131.0, 130.5, 130.4, 129.5, 128.6, 128.3, 125.3, 124.1, 118.9, 118.0, 36.5, 26.8, 20.9. IR (KBr) ν: 3356, 1641, 1548, 1466, 1400, 1309, 1253 cm−1. HR-MS (ESI) calcd for C32H27N2O3 [M + H]+ 487.2022, found 487.2031.
N3,N11-Dipropyl-14-phenyl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7a): White solid. Yield 87.8%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.74 (d, J = 8.9 Hz, 2H, H-1, 13), 8.57 (t, J = 5.5 Hz, 2H, CONH × 2), 8.44 (s, 2H, H-4, 10), 8.06–8.00 (m, 4H), 7.69–7.55 (m, 4H), 7.15 (t, J = 7.6 Hz, 2H, H-3′, 5′), 6.98 (t, J = 7.5 Hz, 1H, H-4′), 6.78 (s, 1H, H-14), 3.30–3.13 (m, 4H, NHCH2CH2CH3 × 2), 1.56 (m, 4H, NHCH2CH2CH3 × 2), 0.91 (t, J = 7.4 Hz, 6H, NHCH2CH2CH3 × 2). IR (KBr) ν: 3272, 2959, 2927, 1638, 1551, 1463, 1250 cm−1. HR-MS (ESI) calcd for C35H33N2O3 [M + H]+ 529.2491, found 529.2498.
N3,N11-Dipropyl-14-(2-fluorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7b): White solid. Yield 91.2%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.57 (t, J = 5.6 Hz, 2H, CONH × 2), 8.47 (d, J = 8.9 Hz, 2H, H-1, 13), 8.46 (s, 2H, H-4, 10), 8.07 (d, J = 9.0 Hz, 2H, H-5, 9), 8.04 (dd, J = 8.9, 1.5 Hz, 2H, H-2, 12), 7.63 (d, J = 8.9 Hz, 2H, H-6, 8), 7.58 (t, J = 7.8 Hz, 1H, H-6′), 7.13–7.05 (m, 2H, H-3′, 5′), 7.04–6.90 (m, 1H, H-4′), 6.89 (s, 1H, H-14), 3.34–3.19 (m, 4H, NHCH2CH2CH3 × 2), 1.56 (h, J = 7.2 Hz, 4H, NHCH2CH2CH3 × 2), 0.91 (t, J = 7.4 Hz, 6H, NHCH2CH2CH3 × 2). IR (KBr) ν: 3270, 2960, 2934, 1632, 1551, 1464, 1252 cm−1. HR-MS (ESI) calcd for C35H32FN2O3 [M + H]+ 547.2397, found 547.2404.
N3,N11-Dipropyl-14-(4-fluorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7c): White solid. Yield 88.7%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.73 (d, J = 9.0 Hz, 2H, H-1, 13), 8.58 (t, J = 5.7 Hz, 2H, CONH × 2), 8.45 (d, J = 1.6 Hz, 2H, H-4, 10), 8.06–8.00 (m, 4H), 7.76–7.54 (m, 4H), 6.98 (t, J = 8.9 Hz, 2H, H-3′, 5′), 6.82 (s, 1H, H-14), 3.35–3.14 (m, 4H, NHCH2CH2CH3 × 2), 1.57 (h, J = 7.3 Hz, 4H, NHCH2CH2CH3 × 2), 0.91 (t, J = 7.4 Hz, 6H, NHCH2CH2CH3 × 2). IR (KBr) ν: 3292, 2962, 2931, 2873, 1634, 1549, 1507, 1462, 1399, 1248 cm−1. HR-MS (ESI) calcd for C35H32FN2O3 [M + H]+ 547.2397, found 547.2391.
N3,N11-Dipropyl-14-(2-chlorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7d): White solid. Yield 84.6%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.67 (d, J = 9.0 Hz, 2H, H-1, 13), 8.58 (t, J = 5.7 Hz, 2H, CONH × 2), 8.47 (d, J = 1.5 Hz, 2H, H-4, 10), 8.09 (d, J = 9.0 Hz, 2H, H-5, 9), 8.05 (dd, J = 8.9, 1.7 Hz, 2H, H-2, 12), 7.65 (d, J = 8.9 Hz, 2H, H-6, 8), 7.54 (d, J = 7.8 Hz, 1H, H-6′), 7.34 (d, J = 6.8 Hz, 1H, H-3′), 7.15 (t, J = 7.0 Hz, 1H, H-5′), 7.07 (t, J = 7.6 Hz, 1H, H-4′), 6.89 (s, 1H, H-14), 3.35–3.16 (m, 4H, NHCH2CH2CH3 × 2), 1.56 (m, 4H, NHCH2CH2CH3 × 2), 0.91 (t, J = 7.4 Hz, 6H, NHCH2CH2CH3 × 2). IR (KBr) ν: 3260, 2960, 2926, 2872, 1639, 1550, 1463, 1398, 1250 cm−1. HR-MS (ESI) calcd for C35H32ClN2O3 [M + H]+ 563.2101, found 563.2112.
N3,N11-Dipropyl-14-(4-chlorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7e): White solid. Yield 82.5%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.72 (d, J = 9.0 Hz, 2H, H-1, 13), 8.58 (t, J = 5.7 Hz, 2H, CONH × 2), 8.46 (d, J = 1.7 Hz, 2H, H-4, 10), 8.06 (d, J = 8.9 Hz, 2H, H-5, 9), 8.03 (dd, J = 9.0, 1.8 Hz, 2H, H-2, 12), 7.64 (d, J = 8.6 Hz, 2H, H-2′, 6′), 7.63 (d, J = 8.9 Hz, 2H, H-6, 8), 7.22 (d, J = 8.6 Hz, 2H, H-3′, 5′), 6.82 (s, 1H, H-14), 3.36–3.19 (m, 4H, NHCH2CH2CH3 × 2), 1.57(h, J = 7.3 Hz, 4H, NHCH2CH2CH3 × 2), 0.91 (t, J = 7.4 Hz, 6H, NHCH2CH2CH3 × 2). IR (KBr) ν: 3302, 2962, 2931, 1636, 1547, 1462, 1399, 1251 cm−1. HR-MS (ESI) calcd for C35H32ClN2O3 [M + H]+ 563.2101, found 563.2096.
N3,N11-Dipropyl-14-(3-nitrophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7f): White solid. Yield 86.3%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.80 (d, J = 9.0 Hz, 2H, H-1, 13), 8.62–8.56 (m, 3H), 8.47 (d, J = 1.6 Hz, 2H, H-4, 10), 8.17–7.99 (m, 5H), 7.87 (dd, J = 8.2, 1.4 Hz, 1H, H-4′), 7.68 (d, J = 8.9 Hz, 2H, H-6, 8), 7.47 (t, J = 8.0 Hz, 1H, H-5′), 7.03 (s, 1H, H-14), 3.38–3.19 (m, 4H, NHCH2CH2CH3 × 2), 1.56 (m, 4H, NHCH2CH2CH3 × 2), 0.91 (t, J = 7.4 Hz, 6H, NHCH2CH2CH3 × 2). IR (KBr) ν: 3243, 2959, 2921, 2868, 1630, 1526, 1462, 1400, 1348, 1319, 1243 cm−1. HR-MS (ESI) calcd for C35H32N3O5 [M + H]+ 574.2342, found 574.2332.
N3,N11-Dipropyl-14-(4-nitrophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7g): White solid. Yield 85.3%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.74 (d, J = 9.0 Hz, 2H, H-1, 13), 8.59 (t, J = 5.7 Hz, 2H, CONH × 2), 8.46 (d, J = 1.7 Hz, 2H, H-4, 10), 8.09 (d, J = 9.0 Hz, 2H, H-3′, 5′), 8.06–8.01(m, 4H), 7.93 (d, J = 8.9 Hz, 2H, H-2′, 6′), 7.67 (d, J = 8.9 Hz, 2H, H-6, 8), 7.00 (s, 1H, H-14), 3.36–3.14(m, 4H, NHCH2CH2CH3 × 2), 1.56 (h, J = 7.3 Hz, 4H, NHCH2CH2CH3 × 2), 0.91 (t, J = 7.4 Hz, 6H, NHCH2CH2CH3 × 2). IR (KBr) ν: 3273, 2966, 2925, 2872, 1639, 1622, 1529, 1463, 1399, 1345, 1250 cm−1. HR-MS (ESI) calcd for C35H32N3O5 [M + H]+ 574.2342, found 574.2350.
N3,N11-Dipropyl-14-(4-methylphenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7h): White solid. Yield 89.9%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.71 (d, J = 9.0 Hz, 2H, H-1, 13), 8.57 (t, J = 5.7Hz, 2H, CONH × 2), 8.44 (d, J = 1.6 Hz, 2H, H-4, 10), 8.03 (d, J = 8.9 Hz, 2H, H-5, 9), 8.01 (dd, J = 9.0, 1.8 Hz, 2H, H-2, 12), 7.62 (d, J = 8.9 Hz, 2H, H-6, 8), 7.49 (d, J = 8.1 Hz, 2H, H-2′, 6′), 6.94 (d, J = 8.0 Hz, 2H, H-3′, 5′), 6.73 (s, 1H, H-14), 3.33–3.16 (m, 4H, NHCH2CH2CH3 × 2), 2.05 (s, 3H, Ar-CH3), 1.57 (h, J = 7.3 Hz, 4H, NHCH2CH2CH3 × 2), 0.91 (t, J = 7.4 Hz, 6H, NHCH2CH2CH3 × 2). IR (KBr) ν: 3289, 2959, 2925, 2856, 1639, 1552, 1511, 1462, 1398, 1317, 1241 cm−1. HR-MS (ESI) calcd for C36H35N2O3 [M + H]+ 543.2648, found 543.2640.
N3,N11-Diisobutyl-14-phenyl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8a): White solid. Yield 84.3%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.74 (d, J = 9.0 Hz, 2H, H-1, 13), 8.58 (t, J = 5.8 Hz, 2H, CONH × 2), 8.45 (d, J = 1.1 Hz, 2H, H-4, 10), 8.07–8.00 (m, 4H), 7.64 (d, J = 8.9 Hz, 2H, H-6, 8), 7.63 (d, J = 7.5 Hz, 2H, H-2′, 6′), 7.15 (t, J = 7.7 Hz, 2H, H-3′, 5′), 6.98 (t, J = 7.3 Hz, 1H, H-4′), 6.78 (s, 1H, H-14), 3.22–3.03 (m, 4H, NHCH2 × 2), 1.94–1.82 (m, 2H, CH(CH3)2 × 2), 0.91 (d, J = 6.7 Hz, 12H, CH(CH3)2 × 2). IR (KBr) ν: 3298, 2958, 2924, 1635, 1550, 1462, 1242 cm−1. HR-MS (ESI) calcd for C37H37N2O3 [M + H]+ 557.2804, found 557.2809.
N3,N11-Diisobutyl-14-(2-fluorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8b): White solid. Yield 82.7%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.57 (t, J = 5.8 Hz, 2H, CONH × 2), 8.50–8.44 (m, 4H), 8.08 (d, J = 9.0 Hz, 2H, H-5, 9), 8.05 (dd, J = 9.0, 1.6 Hz, 2H, H-2, 12), 7.63 (d, J = 8.9 Hz, 2H, H-6, 8), 7.58 (t, J = 7.8 Hz, 1H, H-6′), 7.17–7.05 (m, 2H, H-3′, 5′), 7.04–6.95 (m, 1H, H-4′), 6.89 (s, 1H, H-14), 3.17–3.09 (m, 4H, NHCH2 × 2), 1.94–1.81 (m, 2H, CH(CH3)2 × 2), 0.91 (d, J = 6.7 Hz, 12H, CH(CH3)2 × 2). IR (KBr) ν: 3295, 2958, 2925, 1637, 1550, 1463, 1251 cm−1. HR-MS (ESI) calcd for C37H36FN2O3 [M + H]+ 575.2710, found 575.2715.
N3,N11-Diisobutyl-14-(4-fluorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8c): White solid. Yield 83.8%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.73 (d, J = 9.0 Hz, 2H, H-1, 13), 8.59 (t, J = 5.8 Hz, 2H, CONH × 2), 8.46 (d, J = 1.6 Hz, 2H, H-4, 10), 8.07–8.00 (m, 4H), 7.76–7.54 (m, 4H), 6.98 (t, J = 8.9 Hz, 2H, H-3′, 5′), 6.82 (s, 1H, H-14), 3.21–3.07 (m, 4H, NHCH2 × 2), 1.94–1.81 (m, 2H, CH(CH3)2 × 2), 0.91 (d, J = 6.7 Hz, 12H, CH(CH3)2 × 2). IR (KBr) ν: 3299, 2958, 2925, 1635, 1549, 1507, 1462, 1399, 1248 cm−1. HR-MS (ESI) calcd for C37H36FN2O3 [M + H]+ 575.2710, found 575.2719.
N3,N11-Diisobutyl-14-(2-chlorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8d): White solid. Yield 87.5%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.66 (d, J = 9.0 Hz, 2H, H-1, 13), 8.59 (t, J = 5.8 Hz, 2H, CONH × 2), 8.47 (d, J = 1.3 Hz, 2H, H-4, 10), 8.12–8.03 (m, 4H), 7.64 (d, J = 8.9 Hz, 2H, H-6, 8), 7.53 (d, J = 7.7 Hz, 1H, H-6′), 7.34 (d, J = 8.0 Hz, 1H, H-3′), 7.14 (t, J = 7.1 Hz, 1H, H-5′), 7.07 (t, J = 6.9 Hz, 1H, H-4′), 6.87 (s, 1H, H-14), 3.13 (t, J = 6.4 Hz, 4H, NHCH2 × 2), 1.94–1.81 (m, 2H, CH(CH3)2 × 2), 0.91 (d, J = 6.7 Hz, 12H, CH(CH3)2 × 2). IR (KBr) ν: 3281, 2960, 2926, 1638, 1547, 1463, 1397, 1249 cm−1. HR-MS (ESI) calcd for C37H36ClN2O3 [M + H]+ 591.2414, found 591.2420.
N3,N11-Diisobutyl-14-(4-chlorophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8e): White solid. Yield 81.6%. m.p. 266–268 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.72 (d, J = 9.0 Hz, 2H, H-1, 13), 8.58 (t, J = 5.8 Hz, 2H, CONH × 2), 8.46 (d, J = 1.7 Hz, 2H, H-4, 10), 8.06 (d, J = 8.9 Hz, 2H, H-5, 9), 8.03 (dd, J = 8.9, 1.8 Hz, 2H, H-2, 12), 7.64 (d, J = 8.6 Hz, 2H, H-2′, 6′), 7.63 (d, J = 8.9 Hz, 2H, H-6, 8), 7.22 (d, J = 8.6 Hz, 2H, H-3′, 5′), 6.82 (s, 1H, H-14), 3.20–3.06 (m, 4H, NHCH2 × 2), 1.93–1.80 (m, 2H, CH(CH3)2 × 2), 0.91 (d, J = 6.7 Hz, 12H, CH(CH3)2 × 2). IR (KBr) ν: 3319, 2958, 2926, 1637, 1546, 1463, 1398, 1250 cm−1. HR-MS (ESI) calcd for C37H36ClN2O3 [M + H]+ 591.2414, found 591.2418.
N3,N11-Diisobutyl-14-(3-nitrophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8f): White solid. Yield 85.1%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.80 (d, J = 9.0 Hz, 2H, H-1, 13), 8.62–8.56 (m, 3H), 8.47 (d, J = 1.6 Hz, 2H, H-4, 10), 8.17–7.99 (m, 5H), 7.87 (dd, J = 8.2, 1.4 Hz, 1H, H-4′), 7.68 (d, J = 8.9 Hz, 2H, H-6, 8), 7.48 (t, J = 8.0 Hz, 1H, H-5′), 7.03 (s, 1H, H-14), 3.23–3.02 (m, 4H, NHCH2 × 2), 1.94–1.81 (m, 2H, CH(CH3)2 × 2), 0.91 (d, J = 6.7 Hz, 12H, CH(CH3)2 × 2). IR (KBr) ν: 3293, 2960, 2926, 1640, 1532, 1462, 1396, 1350, 1246 cm−1. HR-MS (ESI) calcd for C37H36N3O5 [M + H]+ 602.2655, found 602.2650.
N3,N11-Diisobutyl-14-(4-nitrophenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8g): White solid. Yield 82.6%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.74 (d, J = 8.9 Hz, 2H, H-1, 13), 8.59 (t, J = 5.7 Hz, 2H, CONH × 2), 8.47 (d, J = 1.6 Hz, 2H, H-4, 10), 8.09 (d, J = 9.0 Hz, 2H, H-3′, 5′), 8.06–8.01(m, 4H), 7.93 (d, J = 8.9 Hz, 2H, H-2′, 6′), 7.67 (d, J = 8.9 Hz, 2H, H-6, 8), 7.00 (s, 1H, H-14), 3.23–3.02 (m, 4H, NHCH2 × 2), 1.94–1.81 (m, 2H, CH(CH3)2 × 2), 0.91 (d, J = 6.7 Hz, 12H, CH(CH3)2 × 2). IR (KBr) ν: 3320, 2958, 2926, 1640, 1546, 1514, 1463, 1346, 1250 cm−1. HR-MS (ESI) calcd for C37H36N3O5 [M + H]+ 602.2655, found 602.2648.
N3,N11-Diisobutyl-14-(4-methylphenyl)-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8h): White solid. Yield 84.6%. m.p. >300 °C. 1H-NMR (400 MHz, DMSO-d6) δ (in ppm): 8.71 (d, J = 9.0 Hz, 2H, H-1, 13), 8.57 (t, J = 5.8 Hz, 2H, CONH × 2), 8.45 (d, J = 1.6 Hz, 2H, H-4, 10), 8.03 (d, J = 8.9 Hz, 2H, H-5, 9), 8.01 (dd, J = 8.9, 1.8 Hz, 2H, H-2, 12), 7.62 (d, J = 8.9 Hz, 2H, H-6, 8), 7.49 (d, J = 8.1 Hz, 2H, H-2′, 6′), 6.94 (d, J = 8.0 Hz, 2H, H-3′, 5′), 6.73 (s, 1H, H-14), 3.23–3.02 (m, 4H, NHCH2 × 2), 2.05 (s, 3H, Ar-CH3), 1.94–1.81 (m, 2H, CH(CH3)2 × 2), 0.91 (d, J = 6.7 Hz, 12H, CH(CH3)2 × 2). IR (KBr) ν: 3340, 2956, 2922, 1641, 1548, 1463, 1397, 1316, 1249 cm−1. HR-MS (ESI) calcd for C38H39N2O3 [M + H]+ 571.2961, found 571.2967.
The 1H-NMR for compound 5a–8h and 13C-NMR for compound 6a–h can be found in Supplementary Materials.
2.2. Cytotoxicity Assay
The synthesized compounds 5a–h, 6a–h, 7a–h and 8a–h were tested for cytotoxicity in four human cancer cell lines, which contained human hepatoma cells (SK-HEP-1, HepG2, SMMC-7721) and acute promyelocytic leukemia cells (NB4), and arsenic trioxide (As2O3) was used as positive control. In addition, the dibenzo[a,j]xanthene derivatives 9a–d (the preparation method of compounds 9a–d according [15], Figure 2), which do not possess functional groups on the 3- and 11-positions, were used as negative standards. The IC50 values of the carboxamide derivatives for antiproliferative activity are listed in Table 1.
Table 1.
IC50 values (μM) of the synthesized compounds for antiproliferative activity.
| Compound | IC50 (μM) for Different Cell Lines a | |||
|---|---|---|---|---|
| HepG2 | SK-HEP-1 | SMMC-7721 | NB4 | |
| 5a | 17.97 ± 0.51 | 20.86 ± 1.12 | 9.05 ± 0.23 | 9.47 ± 0.35 |
| 5b | >50 | 24.06 ± 2.06 | 20.5 ± 1.77 | >50 |
| 5c | 9.68 ± 0.35 | 9.73 ± 0.53 | 11.76 ± 0.87 | 8.25 ± 0.34 |
| 5d | >50 | 26.24 ± 2.87 | 10.47 ± 0.76 | >50 |
| 5e | 11.58 ± 1.05 | 9.82 ± 0.19 | 8.01 ± 0.39 | 8.12 ± 0.23 |
| 5f | >50 | 40.32 ± 1.77 | 29.49 ± 2.50 | >50 |
| 5g | >50 | 32.85 ± 3.13 | >50 | >50 |
| 5h | 36.16 ± 3.91 | 17.19 ± 0.89 | 9.84 ± 0.43 | 8.73 ± 0.53 |
| 6a | 20.31 ± 2.09 | >50 | 14.52 ± 1.83 | 10.32 ± 1.02 |
| 6b | 30.56 ± 2.98 | >50 | 21.23 ± 2.41 | 14.54 ± 1.98 |
| 6c | 6.12 ± 0.25 | 12.23 ± 1.26 | 7.32 ± 0.49 | 0.52 ± 0.032 |
| 6d | 9.21 ± 0.88 | >50 | 40.5 ± 2.96 | 11.6 ± 1.78 |
| 6e | 6.32 ± 0.30 | 14.61 ± 0.96 | 8.15 ± 0.46 | 0.76 ± 0.041 |
| 6f | 40.14 ± 3.21 | >50 | 22.15 ± 1.46 | 23.34 ± 1.57 |
| 6g | 34.22 ± 3.11 | >50 | 14.63 ± 2.83 | 9.12 ± 0.37 |
| 6h | 8.76 ± 0.47 | 20.17 ± 1.19 | 9.54 ± 0.31 | 1.63 ± 0.041 |
| 7a | 30.21 ± 2.88 | >50 | 18.86 ± 1.70 | 23.32 ± 2.42 |
| 7b | 35.67 ± 2.09 | >50 | 41.31 ± 2.11 | 29.76 ± 2.80 |
| 7c | 9.54 ± 0.31 | 12.78 ± 0.14 | 7.43 ± 0.36 | 7.88 ± 0.16 |
| 7d | 11.79 ± 0.23 | >50 | >50 | 12.61 ± 0.82 |
| 7e | 8.95 ± 0.31 | 15.67 ± 0.15 | 12.20 ± 0.89 | 13.83 ± 0.37 |
| 7f | >50 | >50 | 34.12 ± 1.55 | 30.88 ± 1.80 |
| 7g | 22.13 ± 1.30 | >50 | 19.73 ± 0.78 | 10.97 ± 0.45 |
| 7h | 9.08 ± 0.21 | 22.56 ± 0.45 | 16.87 ± 1.38 | 8.87 ± 0.35 |
| 8a | 32.34 ± 1.25 | >50 | 19.76 ± 0.83 | 27.78 ± 2.61 |
| 8b | 40.45 ± 2.52 | >50 | >50 | 32.48 ± 1.01 |
| 8c | 10.50 ± 0.35 | 13.56 ± 0.23 | 15.06 ± 0.88 | 9.85 ± 0.58 |
| 8d | 12.72 ± 0.91 | >50 | >50 | 13.76 ± 1.17 |
| 8e | 11.34 ± 0.50 | 17.75 ± 0.35 | 13.30 ± 0.86 | 15.34 ± 1.56 |
| 8f | >50 | >50 | >50 | 32.65 ± 2.21 |
| 8g | 25.65 ± 1.92 | >50 | 21.80 ± 1.53 | 12.83 ± 0.59 |
| 8h | 10.37 ± 0.49 | 23.67 ± 0.34 | 17.89 ± 1.33 | 10.01 ± 0.41 |
| 9a–d | > 50 | > 50 | > 50 | > 50 |
| As2O3 | 5.92 ± 0.21 | 6.23 ± 0.32 | 9.43 ± 0.50 | 5.31 ± 0.22 |
a Values are means ± standard deviation from three independent experiments.
3. Discussion
3.1. Synthesis of Target Compounds
Compound 3 was mixed with Br2 in acetic acid at room temperature, and then the mixture was refluxed for 3 h during which period three portions of Sn were added; compound 6-bromo-2-naphthol (yield 75%) was obtained by substitution and reduction reactions of compound 3 [12,13]. The mixture of 6-bromo-2-naphthol and CuCN in DMF was heated at 160–170 °C for 4 h under nitrogen atmosphere to give the compound 6-cyano-2-naphthol (yield 83%) [14]. The latter was refluxed for 8 h in a solvent of 10% hydrochloric acid, the newly formed precipitate was filtered and washed with water and EtOAc successively, and then dried to achieve the compound 6-hydroxy-2-naphthalenecarboxylic acid (yield 75%) [12]. A mixture of the naphthalenecarboxylic acid (20 mmol), and an appropriate amount of arylaldehyde (10.5 mmol), glacial acetic acid (20 mL) and concentrated sulfuric acid (1 mL) was stirred at room temperature for 10 min, and then refluxed for 0.5–2 h to afford compounds 4a–h (yield 82–88%) [12].
Compounds 4a–h were firstly converted into the corresponding acyl chlorides (intermediates), and then the intermediates were converted into corresponding dicarboxamides 5a–h with excessive gaseous NH3; they are easy to purify because they have little solubility in CHCl3 which was used as the solvent, and the reactants are soluble. The acyl chlorides were converted into corresponding dicarboxamides 6a–h in the presence of excessive methylamine in aqueous solution. The principle for the purification of compounds 6a–h is the same as that for compounds 5a–h. Target compounds 7a–h were achieved via the reaction of the acyl chlorides with propylamine at room temperature for 2–3 h, and the purification process was also the same as that for compounds 5a–h, but with less solvent CHCl3, in order to improve the yield of the product. This is because the solubility of 7a–h in chloroform is larger than that of 5a–h in chloroform, and the increased liposolubility is due to the introduction of the propyl group to the N atom in the molecule. Variation of the above solubility is better reflected by compounds 8a–h with an isobutyl group on the N atom in the molecule, which can be completely dissolved in the reaction mixture. The processing of compounds 8a–h was completely different from that of the previous samples. The reaction solution was washed with brine and distilled water successively, and then the organic layer was dried with MgSO4, filtered and evaporated to give the crude product, which was recrystallized in petroleum ether-EtOAc to obtain compounds 8a–h (Scheme 1).
The structures of synthesized compounds 5a–h, 6a–h, 7a–h and 8a–h were confirmed by 1H-NMR, HRMS and IR spectra, and 6a–h were further confirmed by 13C-NMR spectra. Since compounds 5a–8h have symmetric structures, the chemical shifts of H-1 and H-13, H-2 and H-12, H-4 and H-10, H-5 and H-9, H-6 and H-8 are identical in the 1H-NMR spectra, respectively. The H-1, H-2 and H-4 intercouple with each other, and the splitting of the peaks in the spectra and the coupling constants are different. H-1 (d, J ≈ 9Hz), H-2 (dd, J1 ≈ 9Hz, J2 < 2 Hz), and H-4 (d, J < 2 Hz) can be distinguished and assigned. The assignment of the H-5 and H-6 is easy to carry out. H-5 and H-6 are in the meta and ortho positions of the oxygen atom (O-7), respectively, and due to the electron-donating effect of the oxygen atom, the chemical shift of H-5 is greater than that of H-6. The hydrogen atoms on the 14-phenyl group are easy to distinguish. In most cases, the chemical shifts of the hydrogen atoms on the 14-phenyl group have less value than those of hydrogen atoms on naphthalene rings, and the chemical shifts of H-2' and H-6' are greater than those of H-3′ and H-5′, without the influence of other substituents. Although H-14 is hydrogen bonded to a saturated carbon atom, which is at the α-position of three phenyl rings, its chemical shift is relatively large (δH-14 > 6.7).
Since the aromatic carboxamide groups of compounds 5a–h cannot rotate freely, in the 1H-NMR spectroscopy, the two hydrogen atoms on the same nitrogen atom exhibit two single peaks with different chemical shifts. The two carboxamide groups of compounds 5a–h are symmetric, so the integral value of each single peak was 2.
3.2. Cytotoxicity Assay
As evidenced by the cytotoxicity data in Table 1, compounds 5a–h, 6a–h, 7a–h and 8a–h show better inhibitory activity than 9a–d, and the latter do not show significant antitumor activity (IC50 > 50 μM). In general, compounds 5a–h and 6a–h had better antitumor activity than 7a–h and 8a–h, especially the derivatives 6c–6e. The latter proved to be the most potent cytotoxic agents to NB4 cancer cells, the IC50 values of which were 0.52 μM and 0.76 μM, respectively, much lower than 5.31 μM of As2O3. In addition, most of the target compounds exhibited more inhibitory activity on the NB4 cancer cell line than on the other cell lines.
The different antitumor activities of the target molecules to four kinds of tumor cell lines may be attributed to multiple factors such as the side chains on the C-3 and C-11 positions, the nature of the substituent at the 14-phenyl group, the nature of the tested cell lines, and so on.
The carboxamide side chains on C-3 and C-11 exert a strong effect on the antitumor activity. It is found that a small substituent on the N atom of the carboxamide side chain is favorable for cytotoxicity, and the order of inhibitory activity against tumor cells is CH3 > (CH2)2CH3 > CH2CH(CH3)2. When a small substituent connects to the N atom, the latter is easier to form a hydrogen bond with the acceptor (6a vs. 7a and 8a, 6c vs. 7c and 8c, 6e vs. 7e and 8e, Table 1).
The antitumor activities of compounds 5a–h and 6a–h cannot be compared simply in terms of strength or weakness (for example, compound 5a shows better inhibitory activity than 6a to the four cell lines, but compound 6c exhibits stronger inhibitory activity than 5c to the HepG2, SMMC-7721 and NB4 cell lines), which may be due to the small size difference between the hydrogen atom and the methyl group on the N atom, so that it does not affect the formation of hydrogen bonds between the amide group and antitumor cells.
A substituent at C-4′ (para) position on the 14-phenyl group of the dicarboxamide derivatives showed better inhibitory activity than the other positions (C-2′ and C-3′) in most cases (compounds 5c–8c vs. compounds 5b–8b, compounds 5e–8e vs. compounds 5d–8d, and compounds 5g–8g vs. compounds 5f–8f, Table 1), which may be because the C-4′ substituent at one end of the molecule can combine with tumor cells more easily. A halogen atom was more potent than other substituents at the C-4′ position of the 14-phenyl ring, which may be due to its smaller size as well as its higher capability of forming hydrogen bonds with the tumor cells.
Several compounds show an obvious inhibition selectivity to tumor cells. For example, compound 6d exhibits anti-proliferative activity toward HepG2 and NB4 cells; however, it does not show cytotoxic activity toward human hepatoma SK-HEP-1 cells (the IC50 value was greater than 50 μM, Table 1). Similar cell selectivity can be found for other compounds such as 5d–6a.
4. Materials and Methods
4.1. General
The melting points of the targeted compounds were determined with an X-6 melting point apparatus (Beijing Tektronix Instrument Co., Ltd., Beijing, China) and were uncorrected. FT-IR spectra were recorded on an Avatar 370 FT-IR spectrometer in the form of KBr pellets (Thermo Nicolet Corporation, Madison, WI, USA). 1H-NMR (300 MHz or 400 MHz) and 13C-NMR (75 MHz) spectra were recorded on a Bruker Avance 300 (or 400) spectrometer (Bruker Company, Billerica, MA, United States) in DMSO-d6 solution, using tetramethylsilane (TMS) as an internal standard. HR-MS were measured on a Waters LCT Premier XE benchtop orthogonal acceleration time-of-flight mass spectrometer (Waters Corporation, Milford, MA, USA). Unless otherwise noted, all common solvent and chemicals were purchased from commercial suppliers and used without further purification.
4.2. General Procedure for the Preparation of 14-Aryl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (5a–h)
The 4 mmol compounds 4, 30 mL of CHCl3 and 1 drop of dimethylformamide were mixed, and then a solution of 4.4 mL (60 mmol) thionyl chloride in CHCl3 (10 mL) was added to the above system, and the mixture was heated to reflux for 4 h. After the solvent and excessive thionyl chloride were removed using a rotary evaporator, and the acyl chlorides can be obtained. The acyl chloride was dissolved in CHCl3 (30 mL) at room temperature, then excessive gaseous NH3 was introduced into the above solution. The mixture was stirred at room temperature for 2–3 h. A white solid formed was filtered, washed with CHCl3 and water successively, and then dried under vacuum to achieve compounds 5a–h.
4.3. General Procedure for the Preparation of N3,N11-Dimethyl-14-aryl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (6a–h)
The acyl chloride (prepared from 4 mmol compound 4 according to the above method) was dissolved in CHCl3 (30 mL), and the solution was dropwise added into 10 mL of a 40% methylamine solution in water. The reaction mixture was then stirred at room temperature, and thin-layer chromatography (TLC) was used to monitor the reactions. After stirring for 2–3 h, a white solid formed was filtered, washed with CHCl3 and water, separately. After vacuum drying, pure compounds 6a–h were obtained.
4.4. General Procedure for the Preparation of N3,N11-Dipropyl-14-aryl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (7a–h)
The acyl chloride (prepared from 4 mmol compound 4 according to the above method) was dissolved in CHCl3 (20 mL), and the solution was dropwise added into a mixed solution of 10 mL CHCl3 and 16 mmol propylamine. The reaction mixture was stirred at room temperature for 2–3 h, and TLC was used to monitor the reactions. The white solid formed was filtered, washed with CHCl3 and water, separately. The solid was recrystallized from petroleum ether–EtOAc (3:1) to give compounds 7a–h.
4.5. General Procedure for the Preparation of N3,N11-Diisobutyl-14-aryl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide (8a–h)
The acyl chloride (prepared from 4 mmol compound 4 according to the above method) was dissolved in CHCl3 (30 mL), and the solution was dropwise added into a mixed solution of 20 mL CHCl3 and 16 mmol isobutylamine. The reaction mixture was stirred at room temperature, and TLC was used to monitor the reaction. Finally the reaction mixture was washed with brine and water successively, and then dried (MgSO4), filtered and concentrated to give the crude products, which were recrystallized from petroleum Ether–EtOAc (6:1) to afford compounds 8a–h.
5. Conclusions
In this paper, 32 N-substituted14-aryl-14H-dibenzo[a,j]xanthene-3,11-dicarboxamide derivatives were designed and synthesized in acceptable overall yields. All compounds were screened for their cytotoxicity against HepG2, SK-HEP-1, SMMC-7721 and NB4 cell lines. The results of the antitumor activity experiments revealed that some of the compounds exhibit promising inhibitory activity, among which compounds 6c–6e are 10-fold and seven-fold more active compared with the positive control As2O3 against NB4 cells, respectively. In summary, the SARs show that unsubstituted and methyl-substituted dicarboxamide derivatives on the N atoms of carboxamide side chains are favorable for cytotoxicity, and the introduction of bigger substitutes including n-propyl and isobutyl groups leads to inferior results. Based on our previous results [7] and the results in this paper, it is concluded that introducing small-sized groups or polar groups to the N atoms of carboxamide side chains in the dicarboxamide derivatives may be beneficial to antitumor activity.
Acknowledgments
This work was supported by the Youth Science Fund of Heilongjiang Province (No.: QC2014C091), UNPYSCT (No.: 324015507) and the China Postdoctoral Science Foundation (No.: 2013M540306). The authors thank Dr. Yi Wang for checking the language.
Supplementary Materials
The following are available online at www.mdpi.com/1420-3049/22/4/517/s1.
Author Contributions
Y.S., Y.Y. and L.W. conceived and designed the experiments; Y.S., H.J., X.D. and G.C. carried out the synthesis of the xanthene derivatives; Y.Y., N.D. and S.G. evaluated the biological activity; Y.S. and B.L. wrote the paper. All authors analyzed the data, and read and approved the final manuscript.
Conflicts of Interest
The authors declare no conflict of interest.
Footnotes
Sample Availability: Samples of the compounds 5a–7h are available from the authors.
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