SUPPORTING MATERIAL

Further studies on anti-invasive chemotypes: an excursion from chalcones to curcuminoids

Bart I. Roman a,c , Tine De Ryck b , Sigrid Verhasselt a , Marc E. Bracke b and Christian V. Stevens a,

a Research Group SynBioC, Department of Sustainable Organic Chemistry and Technology, Faculty of Bioscience Engineering, Coupure Links 653, B-9000 Ghent, Belgium;b Laboratory of Experimental Cancer Research, Department of Radiation Oncology and Experimental Cancer Research, Ghent University, De Pintelaan 185, B-9000 Ghent, Belgium;c B.I.R. is a postdoctoral fellow of the Research Foundation – Flanders (FWO – Vlaanderen)

Contents

1. Experimental details..............................................................................................p. 2

1.1 Synthesis.....................................................................................................................p. 2

1.2 Biology.......................................................................................................................p. 7

2. 1H, 13C and 19F NMR spectra of characterized molecules..................................p. 8

3. References...............................................................................................................p. 23

Corresponding author. Tel.: +32-9-264-5957; fax: +32-9-264-6221; e-mail: chris.stevens@ugent.be; URL: http://www.ugent.be/bw/doct/en.

1

1. Experimental part

1.1. Synthesis

All reagents were purchased from commercial sources (Aldrich, Acros, TCI) and used without

further purification. Solvents were purchased from commercial sources (Aldrich) and employed as is.

Crude reaction mixtures were analyzed on LC/MS/UV. Thin layer chromatography was carried out on

silica gel 60F254 plates (Merck).

High resolution 1H (400 MHz), 13C (100 MHz) and 19F (376 MHz) magnetic resonance (NMR)

spectra were recorded on a Bruker Avance III 400 FT NMR spectrometer in deuterated solvents.

Chemical shifts are reported using TMS and/or CFCl3 as an internal reference, unless otherwise

indicated. Peak assignments were obtained with the aid of DEPT, HSQC, HMBC and COSY spectra.

Attenuated total reflection (ATR) IR spectra were recorded on a Perkin Elmer Spectrum BX

spectrometer, equipped with a ZnSe-crystal, at room temperature. Low-resolution mass spectra were

recorded on an Agilent Technologies 1100 series VL mass spectrometer (ESI, 70 eV). HRMS analyses

were conducted using an HPLC coupled to an Agilent Technologies 6210 series time-of-flight mass

spectrometer equipped with an ESI/APCI-multimode source. Melting points were measured on a

Kofler bench (WME Heizbank, Wagner & Munz) and are uncorrected.

Preparation of (2E,4E)-1,5-diarylpenta-2,4-dien-1-ones

A solution of the appropriate acetophenone (15.89 mmol, 1 equiv.) and 0.1 g LiOH.H2O in 20 mL

of EtOH was stirred at room temperature for 10 minutes. Afterward, 2 mL of a cinnamaldehyde

(15.89 mmol, 1 equiv.) was added drop-wise. The solution was stirred at room temperature for 3 days.

The resulting slurry was filtered; the residue was washed with EtOH and water, and finally

recrystallized from EtOH.

(2E,4E)-1,5-Diphenylpenta-2,4-dien-1-one 9

Yield: 89%, as yellow crystals; mp: 109.5 °C; IR (ATR, cm -1): max: 661, 692, 776, 995, 1018, 1245,

1576, 1586, 1599 (C=C), 1652 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 6.95-7.12 (m, 3H, H, H,

H), 7.31 (br. t, 1H, Jvic= 7.2 Hz, H4), 7.37 (br. t, 2H, Jvic= 7.2 Hz, H3, H5), 7.45-7.52 (m, 4H, H3’, H5’

and H2, H6), 7.53-7.58 (m, 1H, H4’), 7.60 (ddd, 1H, Jtrans= 14.9 Hz, Jvic= 7.2 Hz, Jallyl= 3.1 Hz, H), 7.97

(br. d, 2H, Jvic= 7.0 Hz, H2’, H6’); 13C NMR (CDCl3, 100 MHz, ppm): 125.2, 126.8 (C, C), 127.2 (C2,

C6), 128.2 (C2’, C6’), 128.4 (C3’, C5’), 128.7 (C3, C5), 129.1 (C4), 132.5 (C4’), 135.9 (C1), 138.0 (C1’),

141.7 (C), 144.6 (C), 190.2 (C=O); ESI-MS (+) m/z: 235.1 ([M+H]+, 100), 236.1 ([M+1+H]+, 18);

ESI-HRMS (+) m/z: Anal. Calcd for C17H15O+ (M+H)+: 235.1117. Found: 235.1116.

2

(2E,4E)-5-Phenyl-1-(p-tolyl)penta-2,4-dien-1-one 10

Yield: 67%, as yellow crystals; mp: 112 °C; IR (ATR, cm -1): max: 682, 724, 814, 998, 1247, 1349,

1579, (C=C), 1605, 1651 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 2.42 (s, 3H, Me), 6.95-7.07 (m,

2H, H, H), 7.10 (d, 1H, Jtrans= 14.9 Hz, H), 7.28 (br. d, 2H, Jvic= 8.1 Hz, H2’, H6’), 7.31 (t, 1H, Jvic= 7.3

Hz, H4), 7.37 (t, 2H, Jvic= 7.3 Hz, H3, H5), 7.50 (br. d, 2H, Jvic= 7.3 Hz, H2, H6), 7.59 (ddd, 1H, Jtrans=

14.9 Hz, Jvic= 7.8 Hz, Jallyl= 2.6 Hz, H), 7.89 (d, 2H, Jvic= 8.1 Hz, H3’, H5’). 13C NMR (CDCl3, 100

MHz, ppm): 21.7 (Me), 125.5 (C), 127.0 (C), 127.3, 128.5, 128.8 (C3, C5 and C2, C6 and C3’, C5’),

129.1 (C4), 129.3 (C2’, C6’), 135.7 (C1’), 136.2 (C1), 141.6 (C), 143.5 (C4’), 144.4 (C), 190.0 (C=O);

ESI-MS (+) m/z: 249.1 ([M+H]+, 100), 250.1 ([M+1+H]+, 20); ESI-HRMS (+) m/z: Anal. Calcd for

C18H17O+ (M+H)+: 249.1274. Found: 249.1279.

(2E,4E)-1-(4-Methoxyphenyl)-5-phenylpenta-2,4-dien-1-one 11

Yield: 70%, as yellow crystals; mp: 77.5 °C; IR (ATR, cm -1): max: 693, 813, 999, 1172, 1248, 1568,

1582, 1597 (C=C), 1651 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 3.88 (s, 3H, OMe), 6.97 (d, 2H,

Jvic= 9.0 Hz, H3’, H5’), 6.99-7.07 (m, 2H, H, H), 7.11 (d, 1H, Jtrans= 14.9 Hz, H), 7.32 (br. t, 1H, Jvic=

7.1 Hz, H4), 7.37 (br. t, 2H, Jvic= 7.1 Hz, H3, H5), 7.50 (br. d, 2H, Jvic= 7.1 Hz, H2, H6), 7.60 (ddd, 1H,

Jtrans= 14.9 Hz, Jvic= 8.0 Hz, Jallyl= 2.4 Hz, H), 8.00 (d, 2H, Jvic= 9.0 Hz, H2’, H6’); 13C NMR (CDCl3,

100 MHz, ppm): 55.5 (OMe), 113.8 (C3’, C5’), 125.3 (C), 127.1 (C4), 127.2, 128.8 (C2, C6 and C3,

C5), 129.1 (C), 130.7 (C2’, C6’), 131.1 (C1’), 136.2 (C1), 141.4 (C), 144.0 (C), 163.3 (C4’), 188.7

(C=O); ESI-MS (+) m/z: 265.1 ([M+H]+, 100), 266.1 ([M+H]+, 20);ESI-HRMS (+) m/z: Anal. Calcd

for C18H17O2+ (M+H)+: 265.1223. Found: 265.1224.

(2E,4E)-1-(4-Chlorophenyl)-5-phenylpenta-2,4-dien-1-one 12

Yield: 64%, as yellow crystals; mp: 143 °C; IR (ATR, cm -1): max: 669, 684, 723, 813, 991, 1570,

1582 (C=C), 1648 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 6.96-7.06 (m, 3H, H, H, H), 7.32 (t,

1H, Jvic= 7.0 Hz, H4), 7.37 (t, 2H, Jvic= 7.0 Hz, H3, H5), 7.45 (br. d, 2H, Jvic= 8.6 Hz, H2’, H6’), 7.49 (br.

d, 2H, Jvic= 7.0 Hz, H2, H6), 7.60 (d, 1H, Jtrans= 15.1 Hz, Jvic= 6.0 Hz, Jallyl= 4.4 Hz, H), 7.91 (d, 2H,

Jvic= 8.6 Hz, H3’, H5’); 13C NMR (CDCl3, 100 MHz, ppm): 124.7 (C), 126.7 (C4), 127.3, 128.8, 128.8

(C2, C6 and C3, C5 and C3’, C5’), 129.3 (C), 129.7 (C2’, C6’), 135.9 (C1), 136.5 (C1’), 139.0 (C4’), 142.3

(C), 145.2 (C), 188.9 (C=O); ESI-MS (+) m/z: 269.1 ([M+H]+, 100), 270.1 ([M+1+H]+, 20), 271.1

([M+2+H]+, 33), 282.3 (27), 325.2 (43); ESI-HRMS (+) m/z: Anal. Calcd for C17H14ClO+ (M+H)+:

269.0728. Found: 269.0722.

(2E,4E)-1-(4-Fluorophenyl)-5-phenylpenta-2,4-dien-1-one 13

Yield: 67%, as yellow crystals; mp: 94 °C; IR (ATR, cm -1): max: 693, 820, 1237, 1505, 1576, 1595

(C=C),1652 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 6.97-7.07 (m, 3H, H, H, H), 7.16 (dd, 2H,

Jvic= 8.8 Hz, JHF= 8.5 Hz, H3’, H5’), 7.33 (br. t, 1H, Jvic= 7.3 Hz, H4), 7.38 (br. t, 2H, Jvic= 7.3 Hz, H3,

H5), 7.50 (dd, 1H, Jvic= 7.3 Hz, Jallyl= 1.6 Hz, H2, H6), 7.61 (ddd, 1H, Jtrans= 14.9 Hz, Jvic= 6.1 Hz, Jallyl=

4.3 Hz, H), 8.01 (dd, 2H, Jvic= 8.8 Hz, JHF= 5.4 Hz, H2’, H6’); 13C NMR (CDCl3, 100 MHz, ppm): 115.7 (d, JCF= 21.9 Hz, C2’, C6’), 124.9 (C), 126.8 (C4), 127.3, 128.9 (C2, C6 and C3, C5), 129.3 (C),

3

131.0 (d, JCF= 9.0 Hz, C2’, C6’), 134.6 (d, JCF= 2.9 Hz, C1’), 136.0 (C1), 142.2 (C), 145.0 (C), 165.5 (d,

JCF= 254.0 Hz, C4’), 188.8 (C=O); 19F NMR (CDCl3, 376 MHz, ppm): -106.36 (tt, 3JHF= 8.5 Hz, 4JHF=

5.4 Hz); ESI-MS (+) m/z: 253.1 ([M+H]+, 100), 254.1 ([M+H]+, 18); ESI-HRMS (+) m/z: Anal. Calcd

for C17H14FO+ (M+H)+: 253.1029. Found: 253.1017.

(2E,4E)-1-(4-Nitrophenyl)-5-phenylpenta-2,4-dien-1-one 14

Yield: 90%, as yellow crystals; mp: 172 °C; IR (ATR, cm-1): max: 690, 998, 1321 (N-O), 1520 (N-

O), 1569, 1597 (C=C), 1653 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 6.99-7.13 (m, 3H, H, H,

H), 7.33-7.43 (m, 3H, H3, H4, H5), 7.52 (dd, 2H, Jvic= 8.1 Hz, Jallyl= 1.3 Hz, H2, H6), 7.64 (ddd, 1H,

Jtrans= 15.0 Hz, Jvic= 9.4 Hz, Jallyl= 0.9 Hz, H), 8.10 (d, 2H, Jvic= 8.9 Hz, H2’, H6’), 8.34 (d, 2H, Jvic= 8.9

Hz, H3’, H5’). 13C NMR (CDCl3, 100 MHz, ppm): 123.8 (C3’, C5’), 124.5 (C),126.5 (C4), 127.5,

129.0, 129.3 (C2, C6 and C3, C5 and C2’, C6’), 129.7 (C), 135.8 (C1), 143.1 (C1’), 143.6 (C), 146.8 (C),

150.0 (C4’), 188.9 (C=O); ESI-MS (+) m/z: 280.1 ([M+H]+, 100), 281.1 ([M+1+H]+, 20), 282.3

([M+2+H]+, 60), 325.2 (28); ESI-HRMS (+) m/z: Anal. Calcd for C17H14NO3+ (M+H)+: 280.0968.

Found: 280.0971.

(2E,4E)-1-(3,4,5-Trimethoxyphenyl)-5-phenylpenta-2,4-dien-1-one 15

Yield: 65%, as yellow crystals; mp: 134 °C; IR (ATR, cm -1): max: 715, 1000, 1127, 1331, 1415,

1570 (C=C), 1650 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 3.94 (s, 3H, C4’OMe), 3.95 (s, 6H,

C3’OMe, C5’OMe), 6.99-7.12 (m, 3H, H, H, H), 7.25 (s, 2H, H2’, H6’), 7.35 (br. t, 1H, Jvic= 7.2 Hz,

H4), 7.38 (br. t, 2H, Jvic= 7.2 Hz, H3, H5), 7.51 (dd, 2H, Jvic= 7.2 Hz, Jallyl= 1.5 Hz, H2, H6), 7.62 (ddd,

1H, Jtrans= 14.9 Hz, Jvic= 7.4 Hz, Jallyl= 2.8 Hz, H); 13C NMR (CDCl3, 100 MHz, ppm): 56.4 (C3’OMe,

C5’OMe), 61.0 (C4’OMe), 105.9 (C2’, C6’), 125.0 (C), 126.9 (C), 127.3 (C3, C5), 128.9 (C2, C6), 129.3

(C4), 133.5 (C1’), 136.1 (C1), 141.9 (C), 142.4 (C4’), 144.7 (C), 153.1 (C3’, C5’), 189.0 (C=O); ESI-MS

(+) m/z: 325.1 ([M+H]+, 100), 326.1 ([M+1+H]+, 22); ESI-HRMS (+) m/z: Anal. Calcd for C20H21O4+

(M+H)+: 325.1434. Found: 325.1423.

(2E,4E)-5-(4-Fluorophenyl)-1-(3,4,5-trimethoxyphenyl)phenylpenta-2,4-dien-1-one 16

Yield: 37%, as yellow crystals; mp: 132 °C; IR (ATR, cm -1): max: 717, 841, 991, 1072, 1123, 1165,

1224 1324, 1343, 1414, 1502, 1585 (C=C), 1650 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 3.93 (s,

3H, C4’OMe), 3.94 (s, 6H, C3’OMe, C5’OMe), 6.91-7.11 (m, 5H, H3, H5, H, H, H), 7.25 (s, 2H, H2’,

H6’), 7,47 (dd, 2H, Jvic= 8.7 Hz, JHF= 5.4 Hz, H2, H6), 7,60 (ddd, 1H, Jtrans= 14.7 Hz, Jvic= 7.9 Hz, Jallyl=

2.4 Hz, H); 13C NMR (CDCl3, 100 MHz, ppm): 56.3 (C3’OMe, C5’OMe), 61.0 (C4’OMe), 105.9 (C2’,

C6’), 116.0 (d, JCF= 21.9 Hz, C3, C5), 125.0 (d, JCF= 1.5 Hz, C), 126.7 (d, JCF= 2.5 Hz, C), 129.0 (d,

JCF= 8.2 Hz, C2, C6), 132.4 (d, JCF= 3.3 Hz, C1), 133.5 (C1’), 140.5 (br. s., C), 142.4 (C4’), 144.5 (C),

153.1 (C3’, C5’), 163.2 (d, JCF= 250.2 Hz, C4), 188.9 (C=O); 19F NMR (CDCl3, 376 MHz, ppm): -111.64 (tt, 3JHF= 8.1 Hz, 4JHF= 5.5 Hz); ESI-MS (+) m/z: 343.1 ([M+H]+, 100), 344.1 ([M+1+H]+, 23);

ESI-HRMS (+) m/z: Anal. Calcd for C20H21O4+ (M+H)+: 343.1340. Found: 343.1328.

4

Preparation of (1E,4E)-1,5-diarylpenta-1,4-dien-3-ones

A solution of 2 g 4-phenylbut-3-en-2-one (13.68 mmol, 1 equiv.) and 0.1 g LiOH.H2O in 20 mL

ethanol was stirred at room temperature for 10 minutes. Afterward, an appropriately substituted

benzaldehyde (13.68 mmol, 1 equiv.) was added drop-wise (or portion-wise). The solution was stirred

at room temperature or 40°C for 3 days. The resulting slurry was filtered; the residue was washed with

EtOH and water, and finally recrystallized from EtOH.

(1E,4E)-1,5-diphenylpenta-1,4-dien-3-one 19

Yield: 77%, as yellow crystals; mp: 99.5 °C; IR (ATR, cm-1): max: 695, 760, 981, 1074, 1096, 1191,

1339, 1593 (C=C), 1649 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 7.09 (d, 2H, Jtrans= 16.0 Hz, H,

H’), 7.39-7.44 (m, 6H, H3, H4, H5 and H3’, H4’, H5’), 7.59-7.66 (m, 4H, H2, H2’ and H6, H6’), 7.75 (d,

2H, Jtrans= 16.0 Hz, H, H'); 13C NMR (CDCl3, 100 MHz, ppm): 125.4 (C, C'), 128.4, 128.9 (C3, C3’,

C5, C5’ and C2, C2’, C6, C6’), 130.5 (C4, C4’), 134.8 (C1, C1’), 143.3 (C, C'), 188.9 (C=O); ESI-MS (+)

m/z: 235.1 ([M+H]+, 100), 236.1 (20); ESI-HRMS (+) m/z: Anal. Calcd for C17H15O+ (M+H)+:

235.1117. Found: 235.1116

(1E,4E)-1-(4-Tolyl)-5-phenylpenta-1,4-dien-3-one 20

Yield: 68%, as yellow crystals; mp: 113 °C; IR (ATR, cm -1): max: 690, 815, 994, 1096, 1337, 1585

(C=C), 1619 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 2.39 (s, 3H, Me), 7.04 (d, 1H, Jtrans= 16.1

Hz, H’), 7.08 (d, 1H, Jtrans= 16.0 Hz, H), 7.22 (br. d, 2H, Jvic= 8.1 Hz, H3’, H5’), 7.39-7.44 (m, 3H, H3,

H4, H5), 7.52 (d, 2H, Jvic= 8.1 Hz, H2’ H6’), 7.59-7.64 (m, 2H, H2, H6), 7.72 (d, 1H, Jtrans= 16.1 Hz, H'),

7.73 (d, 1H, Jtrans= 16.0 Hz, H); 13C NMR (CDCl3, 100 MHz, ppm): 21.5 (Me), 124.5 (C'), 125.5

(C), 128.4, 128.4, 129.0, 129.7 (C2, C6 and C3, C5 and C2’, C6’ and C3’, C5’), 130.4 (C4), 132.1 (C1’),

134.9 (C1), 141.1 (C4’), 143.1 (C'), 143.4 (C), 189.0 (C=O); ESI-MS (+) m/z: 249.1 ([M+H]+, 100),

250.1 (20); ESI-HRMS (+) m/z: Anal. Calcd for C18H17O+ (M+H)+: 249.1274, Found: 249.1283.

(1E,4E)-1-(4-Methoxyphenyl)-5-phenylpenta-1,4-dien-3-one 21

Yield: 88%, as yellow crystals; mp: 97 °C; IR (ATR, cm-1): max: 692, 758, 812, 822, 991, 1093, 1175,

1253, 1510, 1568, 1587 (C=C), 1601, 1617 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 3.85 (s, 3H,

OMe), 6.93 (d, 2H, Jvic= 8.6 Hz, H3’, H5’), 6.96 (d, 1H, Jtrans= 15.9 Hz, H’), 7.08 (d, 1H, Jtrans= 15.9 Hz,

H), 7.38-7.44 (m, 3H, H3, H4, H5), 7.60-7.63 (m, 2H, H2, H6), 7.57 (d, 2H, Jvic= 8.6 Hz, H2’ H6’), 7.72

(d, 1H, Jtrans= 15.9 Hz, H'), 7.73 (d, 1H, Jtrans= 15.9 Hz, H), 13C NMR (CDCl3, 100 MHz, ppm): 55.4

(OMe), 114.5 (C3’, C5’), 123.3 (C'), 125.6 (C), 127.5 (C1’), 128.3, 128.9, 130.2 (C2, C6 and C3, C5 and

C2’, C6’), 130.4 (C4), 134.9 (C1), 142.8 (C'), 143.2 (C), 161.7 (C4’), 188.9 (C=O); ESI-MS (+) m/z:

265.1 ([M+H]+, 100), 266.1 (20); ESI-HRMS (+) m/z: Anal. Calcd for C18H17O2+ (M+H)+: 265.1223.

Found: 265.1219.

5

(1E,4E)-1-(4-Chlorophenyl)-5-phenylpenta-1,4-dien-3-one 22

Yield: 87%, as yellow crystals; mp: 130 °C; IR (ATR, cm -1): max: 692, 757, 817, 981, 1087, 1191,

1330, 1490, 1590 (C=C), 1651 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 7.06 (d, 1H, Jtrans= 16.0

Hz, H), 7.06 (d, 1H, Jtrans= 16.0 Hz, H’), 7.39 (H3’, H5’), 7.41-7.45 (m, 3H, H3, H4, H5), 7.55 (H2’, H6’),

7.59-7.65 (m, 2H, H2, H6), 7.69 (d, 1H, Jtrans= 16.0 Hz, H), 7.74 (d, 1H, Jtrans= 16.0 Hz, H'), 13C NMR

(CDCl3, 100 MHz, ppm): 125.4 (C), 125.7 (C'), 128.4, 129.0, 129.3, 129.5 (C3, C5 and C3’, C5’ and

C2, C6 and C2’, C6’), 130.6 (C4), 133.3, 134.7, 136.4 (C1, C1’, C4’), 141.8 (C), 143.6 (C'), 188.6 (C=O);

ESI-MS (+) m/z: 269.0 ([M+H]+, 100), 270.0 ([M+1+H]+, 16), 271.0 ([M+2+H]+, 34), 272.0

([M+3+H]+, 5); ESI-HRMS (+) m/z: Anal. Calcd for C17H15O+ (M+H)+: 269.0728. Found: 269.0731.

(1E,4E)-1-(4-Fluorophenyl)-5-phenylpenta-1,4-dien-3-one 23

Yield: 89%, as yellow crystals; mp: 112 °C; IR (ATR, cm -1): max: 696, 756, 981, 1095, 1157, 1228,

1330, 1506, 1597 (C=C), 1651 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 7.01 (d, 1H, Jtrans= 15.9

Hz, H’), 7.07 (d, 1H, Jtrans= 15.9 Hz, H), 7.11 (t, 2H, Jvic= 8.6 Hz, H3’, H5’), 7.38-7.47 (m, 3H, H3, H4,

H5), 7.54-7.66 (m, 4H, H2’, H6’, H2, H6), 7.70 (d, 1H, Jtrans= 15.9 Hz, H), 7.74 (d, 1H, Jtrans= 15.9 Hz,

H’), 13C NMR (CDCl3, 100 MHz, ppm): 116.1 (d, JCF= 22.0 Hz, C3’, C5’), 125.1 (d, JCF= 2.2 Hz, C’),

125.4 (C), 128.4 (C2, C6), 129.0 (C3, C5), 130.3 (d, JCF= 8.4 Hz, C2’, C6’), 130.6 (C4), 131.1 (d, JCF= 3.5

Hz, C1’), 134.8 (C1) 142.0 (C), 143.4 (C’), 164.0 (d, JCF= 251.7 Hz, C4’), 188.7 (C=O); 19F NMR

(CDCl3, 100 MHz, ppm): -109.64 (tt, 3JHF= 8.4 Hz, 4JHF= 5.4 Hz); ESI-MS (+) m/z: 253.1 ([M+H]+,

100), 254.1 (17); ESI-HRMS (+) m/z: Anal. Calcd for C17H15O+ (M+H)+: 253.1023. Found: 253.1019.

(1E,4E)-1-(4-Nitrophenyl)-5-phenylpenta-1,4-dien-3-one 24

Yield: 90%, as yellow crystals; mp: 152 °C; IR (ATR, cm -1): max: 699, 763, 983, 1191, 1337 (N-O),

1515 (N-O), 1586 (C=C), 1650 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 7.07 (d, 1H, Jtrans= 16.1

Hz, H), 7.20 (d, 1H, Jtrans= 16.0 Hz, H’), 7.41-7.46 (m, 3H, H3, H4, H5), 7.60-7.66 (m, 2H, H2, H6),

7.75 (d, 1H, Jtrans= 16.1 Hz, H), 7.76 (d, 2H, Jvic= 8.7 Hz, H2’, H6’), 7.78 (d, 1H, Jtrans= 16.0 Hz, H'),

8.28 (d, 2H, Jvic= 8.7 Hz, H3’, H5’); 13C NMR (CDCl3, 100 MHz, ppm): 124.2 (C3’, C5’), 125.2 (C),

128.5 (C2, C6), 128.7 (C'), 128.9 (C2’, C6’), 129.1 (C3, C5), 130.9 (C4), 134.5 (C1), 140.1 (C), 141.0

(C1’), 144.5 (C'), 148.6 (C4’), 188.2 (C=O); ESI-MS (+) m/z: 280.0 ([M+H]+, 100), 281.0 ([M+1+H]+,

20), 296.3 (19), 336.1 (18); ESI-HRMS (+) m/z: Anal. Calcd for C17H14NO3+ (M+H)+: 280.0968.

Found: 280.0961.

(1E,4E)-5-(2,4,6-Trimethoxyphenyl)-1-phenylpenta-1,4-dien-3-one 25

Yield: 87%, as yellow crystals; mp: 126 °C; IR (ATR, cm-1): max:697, 822, 1000, 1098, 1204, 1320,

1447, 1556 (C=C), 1606 (C=O); 1H NMR (CDCl3, 400 MHz, ppm): 3.84 (s, 3H, C4’OMe), 3.89 (s,

6H, C2’OMe, C6’OMe), 6.14 (s, 2H, H3’, H5’), 7.08 (d, 1H, Jtrans= 15.9 Hz, H’), 7.30-7.45 (m, 4H, Jvic=

7.6 Hz, H3, H4, H5, H), 7.58 (dd, 2H, Jvic= 7.6 Hz, Jallyl= 1.7 Hz, H2, H6), 7.66 (d, 1H, Jtrans= 15.9 Hz,

H’), 8.14 (d, 1H, Jtrans= 16.2 Hz, H); 13C NMR (CDCl3, 100 MHz, ppm): 55.4 (C4’OMe), 55.8

(C2’OMe, C6’OMe), 91.0 (C3’, C5’), 106.9 (C1’), 126.3, 126.5 (C, C'), 128.2 (C3, C5), 128.8 (C2, C6),

129.8 (C4), 134.6 (C’), 135.7 (C1), 141.6 (C), 161.8 (C2’, C6’), 163.2 (C4’), 190.6 (C=O); ESI-MS (+)

6

m/z: 325.2 ([M+H]+, 100), 326.2 (22); ESI-HRMS (+) m/z: Anal. Calcd for C20H21O4+ (M+H)+:

325.1434. Found: 325.1428.

Synthesis of (1E,4E,6E)-1,7-diphenylhepta-1,4,6-trien-3-one 26

A solution of 2 g 4-phenylbut-3-en-2-one (13.68 mmol, 1 equiv.) and 0.1 g LiOH.H2O in 20 mL of

EtOH was stirred at room temperature for 10 minutes. Afterwards, 1.7 mL of cinnamaldehyde (13.68

mmol, 1 equiv.) was added drop-wise. The solution was stirred at room temperature for 3 days. The

resulting slurry was filtered; the residue was washed with EtOH and water, and finally recrystallized

from EtOAc (40% yield, as yellow crystals).

(1E,4E,6E)-1,7-Diphenylhepta-1,4,6-trien-3-one 26

Yield: 40%, as yellow crystals (from EtOAc); mp: 109 °C (lit. mp 110-111 °C)1; IR (ATR, cm-1): max:

1578, 1599 (C=C), 1664 (C=O), ESI-MS (+) m/z: 261.0 ([M+H]+, 100); ESI-HRMS (+) m/z: Anal.

Calcd for C19H17O+ (M+H)+: 261.1274. Found: 261.1267.

1.2. Biology

Chick heart invasion assay

The assay is based on the confrontation between cancer cell aggregates and embryonic chick heart

fragments in organ culture. A detailed protocol for this assay has been published previously.15

In short, 9-day old precultured heart fragments (PHFs) with a diameter of 0.4 mm were confronted

with aggregates of MCF-7/6 cells with a diameter of 0.2 mm. and incubated at 37 °C on top of semi-

solid agar overnight. Next, the pairs were cultured in suspension for 8 days in a 5 mL Erlenmeyer flask

containing 1.5 mL of culture medium (37 °C, Gyrorotary shaker set to 120 rpm, 10% CO2).

Thereupon, the cultures were fixed in Bouin-Hollande’s solution, embedded in paraffin, serially

sectioned (8 µm) and stained with hematoxylin and eosin.

Compounds were tested in duplo and added to the culture medium as DMSO solutions. Vehicle

cultures (negative control, DMSO) yielded a grade III or IV result. Further information on this assay is

available in the relevant parts of the manuscript.

7

2. 1H, 13C and 19F NMR spectra of characterized molecules

(2E,4E)-1,5-Diphenylpenta-2,4-dien-1-one 9

8

(2E,4E)-5-Phenyl-1-(p-tolyl)penta-2,4-dien-1-one 10

9

(2E,4E)-1-(4-Methoxyphenyl)-5-phenylpenta-2,4-dien-1-one 11

10

(2E,4E)-1-(4-Chlorophenyl)-5-phenylpenta-2,4-dien-1-one 12

11

(2E,4E)-1-(4-Fluorophenyl)-5-phenylpenta-2,4-dien-1-one 13

12

(2E,4E)-1-(4-Nitrophenyl)-5-phenylpenta-2,4-dien-1-one 14

13

(2E,4E)-5-Phenyl-1-(3,4,5-trimethoxyphenyl)penta-2,4-dien-1-one 15

14

(2E,4E)-5-(4-Fluorophenyl)-1-(3,4,5-trimethoxyphenyl)phenylpenta-2,4-dien-1-one 16

15

(1E,4E)-1,5-Diphenylpenta-1,4-dien-3-one 19

16

(1E,4E)-1-(4-Tolyl)-5-phenylpenta-1,4-dien-3-one 20

17

(1E,4E)-1-(4-Methoxyphenyl)-5-phenylpenta-1,4-dien-3-one 21

18

(1E,4E)-1-(4-Chlorophenyl)-5-phenylpenta-1,4-dien-3-one 22

19

(1E,4E)-1-(4-Fluorophenyl)-4-phenylpenta-1,4-dien-3-one 23

20

(1E,4E)-1-(4-Nitro)-5-phenylpenta-1,4-dien-3-one 24

21

(1E,4E)-1-(2,4,6-Trimethoxyphenyl)-5-phenylpenta-1,4-dien-3-one 25

22

References

1. Thiele, G.; Streitwieser, A. J. Am. Chem. Soc. 1994, 116, 446.

23