Synthetic procedures
4.1.1. Synthesis of :1,8-Dichloroanthracene (3), 4,5-Dichloro-9,10-dihydro-9,10-ethanoanthracene-11-carbaldehyde (4), 1,8-Dichloro-9,10-dihydro-9,10-ethanoanthracene-11-carbaldehyde (5), 1-(4,5-Dichloro-9,10-dihydro-9,10-ethanoanthracen-11-yl)-N-methylmethanamine (6) and 1-(1,8-Dichloro-9,10-dihydro-9,10-ethanoanthracen-11-yl)-N-methylmethanamine (7).
The compounds 3-7 were synthesized according to (15, 22) and their characterizations were also recorded in supplementary information.
4.1.2. Synthesis of Z-Ethyl 3-(1,8-dichloro-9,10-dihydro-9,10-ethanoanthracen-11-yl)propenoate (8) and E-Ethyl 3-(1,8-dichloro-9,10-dihydro-9,10-ethanoanthracen-11-yl)propenoate (9)
(Carbethoxymethylene)triphenylphosphorane (2 g, 5.75 mmol) was added to a solution of the aldehyde 5 (1.2 g, 4 mmol) in (36 mL ) CH2Cl2. The reaction mixture was stirred at room temperature for 5 h. The solvent was removed and the residue was purified via flash column chromatography on silica gel using (Ethyl acetateA/Petroleum ether, 1:5) to afford separable isomers 8 and 9 (1.45 g, 97 %) in ratio of 1:2 respectively as yellow oil.
Compound 8: IR (KBr): ν = 3066, 2927, 2860, 1716, 1641, 1575, 1456, 1190, 1029, 771, 759,594 cm-1; 1H NMR (CDCl3, 400 MHz): δ = 1.20 (t; J = 7.3, 3H, -O-CH2-CH3), 1.24-1.29 (m; 1H, H-12) 2.11-2.14 (m; 1H, H-12), 3.76 (m; 1H, H-11), 4.08 (q; J = 7.3, 2H, -O-CH2-CH3), 4.18 (d; J = 2.9, 1H, H-10), 5.29 (t; J = 2.9, 1H, H-9), 5.47 (dd; J = 11.3, 9.5, 1H, -CH=CH-COO-), 5.61 (d; J = 11.7, 1H, -CH=CH-COO-), 6.96-7.19 (m; 6 H, ArH) ppm; 13C NMR (CDCl3, 100 MHz): δ =14.2, 33.1, 36.5, 36.9, 49.7, 60.0, 119.6, 122.1, 123.7, 126.6, 126.8, 127.0, 129.3, 129.5, 139.5, 140.3, 142.3, 145.2, 152.2, 166.0 ppm; MS (EI): m/z (%) = 372 (10) [M+], 367 (5), 248 (65), 246 (100), 176 (18), 131 (5), 69 (12); HRMS (EI): Calcd. For C21H18O2Cl2 [M+] 372.0684, Found 372.0683.
Compound 9: IR (KBr): ν = 3066, 2979, 2935, 2898, 1718, 1650, 1577, 1456, 1446, 1369, 1271, 1180, 1039, 985, 769, 740, 703, 590 cm-1; 1H NMR (CDCl3, 400 MHz): δ = 1.22 (t; J = 7.3, 3H, -O-CH2-CH3), 1.22-1.24 (m; 1H, H-12) 2.04-2.07 (m; 1H, H-12), 2.71 (m;1H, H-11), 4.12 (q; J = 7.3, 2H, -O-CH2-CH3), 4.20 (d; J = 2.2, 1H, H-10), 5.35 (t; J = 2.5, 1H, H-9), 5.75 (d; J = 15.4, 1H, -CH=CH-COO-), 6.36 (dd; J = 15.4, 9.5, 1H, -CH=CH-COO-), 7.04-7.24 (m; 6 H, ArH) ppm; 13C NMR (CDCl3, 100 MHz): δ = 14.2, 32.0, 36.8, 40.9, 49.7, 60.3, 121.5, 121.9, 123.8, 126.7, 126.9, 127.0, 127.1, 129.3, 129.8, 139.5, 140.2, 141.6, 145.1, 150.5, 166.2 ppm; MS (EI): m/z (%) = 372 (41) [M+], 367 (19), 248 (62), 246 (100), 176 (29); HRMS (EI): Calcd. For C21H18O2Cl2 [M+] 372.0684, Found 372.0683.
4.1.3. Synthesis of Ethyl 3-(1,8-dichloro-9,10-dihydro-9,10-ethanoanthracen-11-yl)propanoate (10)
In a two-necked round-bottomed flask (0.37 g of 10% Pd/C) was wetted with ethanol and the flask was evacuated, and flushed with hydrogen two times, then a solution of (1.3 g, 3.5 mmol) unsaturated ester 8 and 9 in (40 mL ) ethanol was added to the reaction mixture. The mixture was stirred for 24 h at room temperature under H2 (balloon). The reaction mixture was filtered through a pad of celite and the solvent was removed in vacuo to afford the corresponding 10 (1.2 g, 92%) as yellow oil.
IR (KBr): ν = 3020, 2933, 2900, 1733, 1460, 1375, 1261, 1176, 1029, 754, 559 cm-1; 1HNMR (CDCl3, 400 MHz): δ = 1.12-1.16 (m; 2H, H-/1), 1.24 (t; J = 7.3, 3H, -O-CH2-CH3), 1.46-1.51 (m; 1H, H-12), 1.86-1.93 (m; 1H, H-11), 1.97-2.04 (m; 1H, H-12), 2.31 (t; J = 8.0, 2H, H-/2), 4.08 (q; J =7.3, 2H, -O-CH2-CH3), 4.15 (d; J = 2.2, 1H, H-10), 5.29 (t; J = 2.5, 1H, H-9), 6.99-7.25 (m; 6 H, ArH) ppm; 13C NMR (CDCl3, 100 MHz): δ =14.1, 31.1, 32.4, 34.3, 37.9, 44.1, 48.7, 60.2, 122.9, 123.2, 123.3, 125.2, 125.4, 125.5, 125.5, 125.8, 140.4, 143.2, 143.7, 144.2, 173.4 ppm; MS (EI): m/z (%) = 374 ([M+], not recorded), 331 (11), 329 (25), 295 (12), 248 (58), 246 (100), 212 (46), 178 (45).
4.1.4. Synthesis of 3-(1,8-Dichloro-9,10-dihydro-9,10-ethanoanthracen-11-yl)propan-1-ol (11)
To a solution of saturated ester 10 (600 mg, 1.6 mmol) in CH2Cl2 (6 mL ), DIBAL (7 mL ) was added. The reaction mixture was stirred for 5 h at room temperature. Then the reaction mixture was quenched with Methanol (1 mL ) followed by the addition of ethyl acetate (30 mL ) and saturated aqueous of NH4Cl (10 mL ). The quenched reaction mixture was filtered through suction funnel and extracted with CH2Cl2 and the combined organic layers were washed with brine, dried over MgSO4, filtered, and concentrated in vacuo. The residue was purified via flash column chromatography on silica gel using ethyl acetate / hexane (1:3) to afford 11 (280 mg, 53 %) as a milky viscous oil.
IR (KBr): ν = 3577, 3336, 2970, 2933, 2860, 1456, 1055, 756, 567cm-1; 1HNMR (CDCl3, 400 MHz): δ = 0.87-0.98 (m; 2H, H-/1), 1.13-1.28 (m; 1H, H-12), 1.53-1.63 (m; 1H, H-11), 1.84-1.93 (m; 2H, H-/2), 1.97-2.08 (m; 1H, H-12), 3.47 (t; J = 6.6, 2H, -CH2OH), 4.04 (d; J = 2.2, 1H, H-10), 4.16 (t; J = 2.5, 1H, H-9), 7.01-7.17 (m; 6 H, ArH) ppm; 13C NMR (CDCl3, 100 MHz): δ = 30.6, 32.2, 34.6, 38.2, 44.3, 48.9, 62.8, 122.9, 123.1, 123.3, 125.2, 125.3, 125.4, 125.5, 125.6, 140.7, 143.3, 143.8, 144.4 ppm; MS (EI): m/z (%) = 332 ([M+], not recorded), 295 (7), 264 (12), 212 (29), 178 (100), 1152 (4).
4.1.5. Synthesis of 3-(1,8-Dichloro-9,10-dihydro-9,10-ethanoanthracen-11-yl)propanal (12)
To a solution of alcohol 11 (250 mg, 0.75 mmol) in CH2Cl2 (6 mL ), PCC (250 mg, 1.2 mmol) was added. The reaction mixture was stirred for 2 h at room temperature. The reaction mixture was concentrated in vacuo and the residue was purified via flash column chromatography on silica gel using ethyl acetate / hexane (1:3) to afford 12 (250 mg, 100 %) as a colorless oil.
IR (KBr): ν = 3020, 2935, 2862, 2812, 1726, 1460, 1172, 1026, 760, 754, 559 cm-1; 1HNMR (CDCl3, 400 MHz): δ = 1.12-1.25 (m; 2H, H-/1), 1.43-1.50 (m; 1H, H-12), 1.84-1.90 (m; 1H, H-11), 1.97-2.04 (m; 1H, H-12), 2.41-2.45 (m, 2H, H-/2), 4.10 (d; J = 2.2, 1H, H-10), 4.25 (t; J = 2.9, 1H, H-9), 7.09-7.25 (m; 6 H, ArH), 9.69 (t; J = 1.4, 1H, CHO) ppm; 13C NMR (CDCl3, 100 MHz): δ = 28.3, 34.5, 37.9, 41.9, 44.1, 48.7, 123.0, 123.2, 123.4, 125.6, 125.5, 125.6, 125.6, 125.9, 140.2, 143.1, 143.7, 144.0, 202.1 ppm; MS (EI): m/z (%) = 330 ([M+], not recorded), 321 (7), 319 (11), 311 (12), 289 (7), 251 (6), 225 (8), 204 (11), 201 (21), 199 (51), 197 (92), 181 (28), 165 (100), 151 (38), 149 (15).
4.1.6. Synthesis of 3-(1,8-Dichloro-9,10-dihydro-9,10-ethanoanthracen-11-yl)-N-methylpropan-1-amine (13)
In a two-necked round-bottom flask (100 mg, 10% Pd/C) was wetted with dichloromethane and the flask was evacuated, flushed with hydrogen two times, then a solution of (110 mg, 0.33 mmol) aldehyde 12 in (5 mL ) methanol was added to the reaction mixture followed by the addition of (0.7 mL , 2 M) solution of methylamine in methanol. The mixture was stirred for 4 h at room temperature under H2 (balloon). The reaction mixture was filtered through a pad of celite and the solvent was removed in vacuo to yield (90 mg, 79 %) of the corresponding amine 13 as white powder, mp: 178 0C.
IR (KBr): ν = 3414, 2935, 2864, 1471, 1399, 1171, 1034, 804, 752, 551, 466 cm-1; 1HNMR (CDCl3, 400 MHz): δ = 0.79-0.87 (m; 2H, H-/1), 1.0-1.10 (m; 2H, H-/2), 1.2 (s; 3H, N-CH3), 1.73-1.79 (m; 1H, H-11), 1.85-1.92 (m; 2H, H-12), 2.68-2.72 (m; 2H, H-/3), 3.18 (d; J = 1.8, 3H, N-CH3), 4.03 (d; J = 2.2, 1H, H-10), 4.13 (t; J = 2.2, 1H, H-9), 6.97-7.02 (m; 3H, ArH), 7.13-7.18 (m; 3H, ArH) ppm; 13C NMR (CDCl3, 100 MHz): δ = 23.8, 32.8, 32.9, 34.4, 37.7, 44.1, 48.5, 49.2, 122.9, 123.3(2x), 125.2, 125.4, 125.5, 125.6, 125.8, 140.3, 143.2, 143.6, 144.0 ppm; MS (EI): m/z (%) = 346 (18) [M+ + H], 336 (19), 335 (31), 334 (75), 332 (100), 326 (5), 318 (11), 298 (11), 286 (5), 284 (9); HRMS (EI): Calcd. For C20H22NCl2 [M+] 346.1129, Found 346.1128.
MTT assay protocol
In vitro antiproliferative activities of the synthesized chlorinated maprotiline analogues 6, 7 and 13 as well intermediates 4 and 5 were demonstrated by determining the IC50 values against three cancer cell lines; the lung carcinoma cell line A549, the hepatocellular carcinoma HepG2 cell line and the colorectal carcinoma HTC-116 cell line (Table2.). Growth inhibitions were measured in 96-well plates. Aliquots of 120 µL of the suspended cells (50,000 mL -1) were given to 60 µL of a serial dilution of the tested compound. After 5 days of incubations, growths were determined the MTT assay. Briefly, 20 μl MTT (5 mg/mL in PBS) was added to each well, and the plate was incubated for 2 h at 37 0C, and 5% CO2-atmosphere in the cell incubator. The supernatants were then discarded and 200 μL of isopropanol/ HCl was added to each well. The absorbance was then read at 550 nm using a microplate reader (Thermo Scientific, USA). The viability of the cells was calculated by dividing the absorbance average of the treated cells by the absorbance average of the control cells multiply 100%. The IC50 values were defined as a concentration that inhibits 50% of cell growth. The activities of the cells were plotted against the concentration of the drugs, and the IC50 values were calculated from the regression curves.