Influence of S-Oxidation on Cytotoxic Activity of Oxathiole-Fused Chalcones

Article abstract of DOI:10.1111/cbdd.12776

Synthesis, in vitro cytotoxic activity, and interaction with tubulin of oxidized, isomeric 1-(5-alkoxybenzo[d][1,3]oxathiol-6-yl)-3-phenylprop-2-en-1-ones and 1-(6-alkoxybenzo[d][1,3]oxathiol-5-yl)-3-phenylprop-2-en-1-ones are described. Most of the compo

Full text of DOI:10.1111/cbdd.12776

Chem Biol Drug Des 2016
Research Article
Influence of S-Oxidation on Cytotoxic Activity of
Oxathiole-Fused Chalcones
Marek T. Konieczny1,*, Anita Buɬakowska2,
Danuta Pirska2, Wojciech Konieczny2, Andrzej
Skladanowski3, Michal Sabisz3,†, Marek
Wojciechowski3 and Krzysztof Lemke4,‡
Received 20 November 2015, revised 12 April 2016 and
accepted for publication 13 April 2016
Chalcones are widely explored as potential lead com-
pounds for new therapeutic agents, and antitumor proper-
ties of the compounds are among the most often studied
(1–17).
1Department of Chemical Technology of Drugs, Medical
University of Gdansk, Gen. J. Hallera Street 107, 80-416
Gdansk, Poland
2Department of Organic Chemistry, Medical University of
Gdansk, 80-416 Gdansk, Poland
We have already reported synthesis and cytotoxic activity of
isomeric oxathiole-fused chalcones 1 (18) and 2 (19), and
their dioxole analogs 3 (18) (Figure 1). It was found that
cytotoxic activity of the compounds depended strongly on
the substituents OR, X, and on the presence of heterocyclic
ring, but was almost insensitive to position of the sulfur
atom, or even its replacement by oxygen (18,19).
3Department of Pharmaceutical Technology and
Biochemistry, Gdansk University of Technology, 80-233
Gdansk, Poland
4Drug Discovery Department, Adamed Group, 05-152
Czosnow, Poland
*Corresponding author: Marek T. Konieczny,
markon@gumed.edu.pl
†Present Addresses: Laboratory of Therapeutic Proteins
and Peptides, Ecole Polytechnique Federale de Lausanne,
CH – 1015, Lausanne, Switzerland
Promising activity of compounds 1 and 2 prompted us to
synthesize their analogs with the sulfur atom oxidized
either to sulfoxide (4, 5) or sulfone (6, 7) (Figure 1). It was
speculated that such modification could increase meta-
bolic stability of the compounds, as oxidation of sulfur
atom to sulfoxide or sulfone constitutes a dominating
metabolic transformation of divalent sulfur compounds
(20–22). Generally, influence of oxidation of sulfur
compounds on their biological activity seems to be unpre-
dictable, as nicely summarized by Prof. A. G. Renwick in
his review on pharmacological and toxicological conse-
quences of sulphoxide oxidation and reduction: “Thus
overall conclusions and generalizations concerning the
effects of changes in redox state on pharmacological or
toxicological activity are probably best avoided” (21,22).
The obtained compounds were tested for their cytotoxic
activity in vitro, and mechanism of the activity was
probed using assays for their interaction with tubulin, and
molecular modeling.
‡Research and Development Department Biovico Ltd. 81-
340 Gdynia Poland
Synthesis, in vitro cytotoxic activity, and interaction
with tubulin of oxidized, isomeric 1-(5-alkoxybenzo[d]
[1,3]oxathiol-6-yl)-3-phenylprop-2-en-1-ones and 1-(6-
alkoxybenzo[d][1,3]oxathiol-5-yl)-3-phenylprop-2-en-1-
ones are described. Most of the compounds
demonstrated cytotoxic activity at submicromolar con-
centrations. It was found that oxidation of sulfur atom
of the oxathiole-fused chalcones strongly influenced
activity of the parent compounds, and that depending
on relative position of the sulfur atom in the molecule,
the activity was either increased or diminished. For
isomers with sulfur atom para to the chalcone
carbonyl group, oxidation led to increase in activity,
while for isomers with sulfur atom meta to the
carbonyl the activity dropped down. It was
demonstrated that the compounds interact with tubu-
lin at the colchicine binding site, and the interaction
was evaluated using molecular modeling. It was
concluded that the observed profound influence of
oxidation of the sulfur atom on cytotoxic activity
cannot be solely related to interaction of the
compounds with tubulin.
Experimental
Chemistry
General
Melting points were determined using a Stuart SMP3
instrument and were uncorrected. Infrared spectra were
obtained from KBr pellets on a Thermo Mattson Satellite
instrument. The 1H NMR spectra were recorded on
Key words: benzoxathiole oxidation, chalcone, cytotoxic
activity, molecular modeling, tubulin interaction
ª 2016 John Wiley & Sons A/S. doi: 10.1111/cbdd.12776
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Konieczny et al.
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Figure 1: Structures of the studied compounds (4–7) and their precursors (1–3).
200 MHz (Varian Gemini) or 500 MHz (Varian Unity Plus)
spectrometers and the chemical shifts were expressed in
J = 16.1 Hz, H-a), 5.65 (d, 1H, J = 11.2 Hz, H-2), 5.19 (d,
1H, J = 11.2 Hz, H-2), 3.86 (s, 3H, OCH3).
d
(ppm) values (s = singlet, d = doublet, t = triplet,
q = quartet, m = multiplet). Elemental analyses were per-
formed using a Carlo-Erba 1108 instrument and results
were within 0.4% of theoretical values. The described
reactions are unoptimized.
EA: Calcd for C17H13ClO4S:, 58.54; H, 3.76; S, 9.19.
Found: 58.31; H, 3.66; S, 8.99.
(E)-1-(5-Methoxy-3-oxidobenzo[d][1,3]oxathiol-6-yl)-3-(4-
methoxyphenyl)prop-2-en-1-one (4c), crystallization solvent:
methanol, a cream solid, yield 85%, mp 201–202 °C.
General procedure for oxidation of chalcones to
their S-monooxido derivatives
IR (KBr, cmꢀ1): 1634, 1602, 1512, 1401, 1255, 1171.
A suitable chalcone (1 mmol) was dissolved in acetic acid
(15 mL), 30% hydrogen peroxide (3 mL) was added and
the mixture was stirred at room temperature for about 2 h.
Icy water was added to the obtained solution and the pre-
cipitated solid was filtered off, washed with water, dried
and crystallized from a suitable solvent, usually methanol
or ethanol.
1H NMR (500 MHz, DMSO-d6): d 7.93 (s, 1H, H-4), 7.71
(d, 2H, J = 8.7 Hz, H-20, H-60), 7.41 (d, 1H, J = 16.1 Hz,
H-b), 7.35 (s, 1H, H-7), 7.14 (d, 1H, J = 16.1 Hz, H-a),
6.99 (d, 2H, J = 8.3 Hz, H-30, H-50), 5.64 (d, 1H,
J = 11.7 Hz, H-2), 5.19 (d, 1H, J = 11.7 Hz, H-2), 3.85 (s,
3H, OCH3), 3.80 (s, 3H, OCH3).
(E)-1-(5-Methoxy-3-oxidobenzo[d][1,3]oxathiol-6-yl)-3-phe-
nylprop-2-en-1-one (4a), crystallization solvent: methanol,
a colorless solid, yield 82%, mp 220–222 °C.
13C NMR (126 MHz, DMSO-d6) d 192.93, 162.26, 153.79,
153.06, 145.73, 136.21, 133.10, 133.07, 131.43, 127.48,
124.90, 115.21, 114.17, 113.25, 111.26, 92.03, 57.43,
56.08.
IR (KBr, cmꢀ1): 3059, 1633, 1475, 1053.
EA: Calcd for C18H16O5S: C, 62.78; H, 4.68; S, 9.31.
Found: C, 62.61; H, 4.65; S, 9.33.
1H NMR (500 MHz, DMSO-d6): d 7.95 (s, 1H, H-4), 7.75
(m, 2H, H-20, H-60), 7.42–7.50 (m, 4H, H-30, H-40, H-50, H-
b), 7.38 (s, 1H, H-7), 7.30 (d, 1H, J = 16.1 Hz, H-a), 5.60
(d, 1H, J = 11.2 Hz, H-2), 5.20 (d, 1H, J = 11.2 Hz, H-2),
3.87 (s, 3H, OCH3).
(E)-3-(3-Hydroxy-4-methoxyphenyl)-1-(5-methoxy-3-oxido-
benzo[d][1,3]oxathiol-6-yl)prop-2-en-1-one (4d), crystalliza-
tion solvent: ethanol, a yellow solid, yield 74%, mp 184–
186 °C.
EA: Calcd for C17H14O4S: C, 64.95; H, 4.49; S, 10.20.
Found: C, 65.33; H, 4.45; S, 10.33.
IR (KBr, cmꢀ1): 3240, 2984, 1653, 1569, 1510, 1400,
1254, 1010, 807.
(E)-3-(3-Chlorophenyl)-1-(5-methoxy-3-oxidobenzo[d][1,3]
oxathiol-6-yl)prop-2-en-1-one (4b), crystallization solvent:
methanol, a cream solid, yield 80%, mp 185–186 °C.
1H NMR (500 MHz, DMSO-d6): d 9.26 (s, 1H, OH), 7.94 (s,
1H, H-4), 7.33 (m, 2H, H-20, H-b), 7.16 (m, 2H, H-60, H-7),
7.02 (d, 1H, J = 16.1 Hz, H-a), 6.97 (d, 1H, J = 8.3 Hz, H-
50), 5.65 (d, 2H, J = 11.2 Hz, H-2), 5.20 (d, 2H,
J = 11.2 Hz, H-2), 3.86 (s, 3H, OCH3), 3.83 (s, 3H, OCH3).
IR (KBr, cmꢀ1): 3065, 1654, 1586, 1471, 1010.
1H NMR (500 MHz, DMSO-d6): d 7.96 (s, 1H, H-4), 7.90
(s, 1H, H-20), 7.74 (d, 1H, J = 7.3 Hz, H-60), 7.44–7.54 (m,
3H, H-40, H-50, H-b), 7.39 (s, 1H, H-7), 7.38 (d, 1H,
13C NMR (126 MHz, DMSO-d6) d 192.67, 153.80, 153.09,
151.28, 147.43, 146.12, 136.17, 133.13, 127.71, 124.64,
2
Chem Biol Drug Des 2016

SAR of Cytotoxic Oxathiole-Fused Chalcones
122.96, 114.95, 113.30, 112.66, 111.29, 92.03, 57.44,
56.34.
J = 7.8 Hz, H-50), 5.64 (d, 1H, J = 11.3 Hz, H-2), 5.19 (d,
1H, J = 11.3 Hz, H-2), 4.06 (s, 2H, OCH2), 3.83 (s, 3H,
OCH3), 1.68 (m, 2H, CH2), 0.89 (t, 3H, J = 6.9 Hz, CH3).
EA: Calcd for C18H16O6S: C, 59.99; H, 4.47; S, 8.90.
Found: C, 59.85;H, 4.38; S, 9.05.
13C NMR (126 MHz, DMSO-d6) d 192.50, 153.71, 152.57,
151.20, 147.41, 145.53, 136.29, 133.37, 127.78, 124.63,
122.79, 114.90, 113.40, 112.65, 112.05, 92.00, 71.31,
56.32, 22.63, 11.12.
(E)-3-(3-Fluoro-4-methoxyphenyl)-1-(5-methoxy-3-oxido-
benzo[d][1,3]oxathiol-6-yl)prop-2-en-1-one (4e), crystalliza-
tion solvent: ethanol, a colorless solid, yield 80%, mp 204–
207 °C.
EA: Calcd for C20H20O6S: C, 61.84; H, 5.19; S, 8.25.
Found: C, 61.69; H, 5.40; S, 8.04.
IR (KBr, cmꢀ1): 2949, 1605, 1521, 1476, 1403, 1289,
1040, 819.
Sodium (E)-2-methoxy-5-(3-(3-oxido-5-propoxybenzo[d][1,3]
oxathiol-6-yl)-3-oxoprop-1-en-1-yl)phenyl phosphate (4h).
The starting (E)-2-methoxy-5-(3-oxo-3-(5-propoxybenzo[d]
[1,3]oxathiol-6-yl)prop-1-en-1-yl)phenyl dihydrogen phos-
phate (1.274 g, 2.81 mmol) was stirred in acetic acid
(25 mL) and hydrogen peroxide (9 mL) for 4 h. The obtained
yellow solid was filtered off, and washed with acetic acid and
next with ethyl ether. The obtained solid was suspended in
cold methanol (5 mL) and 1 N solution of sodium hydroxide
in methanol (5 mL) was added, to give an orange solution.
The solution was concentrated on rotary evaporator, and
ethanol (3 mL) was added to the residue. The precipitate
was filtered off and washed with ethanol to give a beige solid,
yield 1.0 g, 70%, mp 205–208 °C.
1H NMR (500 MHz, DMSO-d6): d 7.95 (s, 1H, H-4), 7.76
(dd, 1H, J1 = 1.9 Hz, J2 = 12.7 Hz H-20), 7.56 (d, 1H,
J = 8.3 Hz, H-60), 7.40 (d, 1H, J = 16.1 Hz, H-b,), 7.36 (s,
1H, H-7), 7.22 (m, 2H, H-50, H-a), 5.65 (d, 1H,
J = 11.2 Hz, H-2), 5.20 (d, 1H, J = 11.2 Hz, H-2), 3.89 (s,
3H, OCH3), 3.86 (s, 3H, OCH3).
13C NMR (126 MHz, DMSO-d6) d 192.94, 153.77, 153.18,
153.11, 151.23, 150.12, 150.04, 144.59, 136.02, 133.19,
128.14, 128.09, 127.61, 127.59, 126.22, 115.96, 115.81,
114.52, 113.28, 111.30, 92.04, 57.44, 56.86.
EA: Calcd for C18H15FO5S: C, 59.66; H, 4.17; S, 8.85.
Found: C, 59.52; H, 4.16; S, 8.84.
IR (KBr, cmꢀ1): 3411, 1655, 1595, 1509, 1270, 1134,
985.
(E)-1-(5-Methoxy-3-oxidobenzo[d][1,3]oxathiol-6-yl)-3-(2,4,6-
trimethoxyphenyl)prop-2-en-1-one (4f), purified on silica gel
column in methylene chloride, followed by crystallization from
ethanol, a yellow solid, yield 50%, mp 209–210 °C.
1H NMR (500 MHz, D2O): d 7.64 (s, 1H, H-4), 7.61 (d, 1H,
J = 2.0 Hz, H-20), 7.33 (d, 1H, J = 16.0 Hz, H-b), 7.23 (s,
1H, H-7), 7.17 (dd, 1H, J1 = 8.5 Hz, J2 = 2.0 Hz, H-60),
6.98 (d, 1H, J = 16.0 Hz, H-a), 6.93 (d, 1H, J = 8.5 Hz,
H-50), 5.58 (d, 1H, J = 11.8 Hz, H-2), 5.15 (d, 1H,
J = 11.8 Hz, H-2), 3.94 (t, 2H, J = 6.4 Hz, OCH2), 3.77
(s, 3H, OCH3), 1.56 (m, 2H, CH2), 0.75 (t, 3H, J = 7.4 Hz,
CH3).
IR (KBr, cmꢀ1): 2944, 1655, 1603, 1561, 1470, 1337,
1203, 1037, 1037, 1009.
1H NMR (500 MHz, DMSO-d6): d 7.92 (s, 1H, H-4), 7.83
(d, 1H, J = 16.1 Hz, H-b), 7.40 (d, 1H, J = 16.1 Hz, H-a),
7.31 (s, 1H, H-7), 6.30 (s, 2H, H-30, H-40), 5.64 (d, 1H,
J = 11.2 Hz H-2), 5.20 (d, 1H, J = 11.2 Hz, H-2,), 3.86 (s,
12H, 4 9 OCH3).
13C NMR (126 MHz, DMSO-d6) d 192.58, 153.71, 152.35,
147.85, 136.79, 132.96, 128.00, 122.97, 115.18, 113.19,
111.93, 111.61, 91.97, 71.25, 55.78, 22.59, 11.07.
EA: Calcd for C20H20O7S: C, 59.40; H, 4.98; S, 7.93.
Found: C, 58.97; H, 5.00; S, 7.80.
EA: Calcd for C20H19Na2O9PS 9 3 H2O: C, 42.41; H,
4.45; S, 5.66. Found: C, 42.67; H, 4.31; S, 5.69.
(E)-3-(3-Hydroxy-4-methoxyphenyl)-1-(3-oxido-5-propoxy-
benzo[d][1,3]oxathiol-6-yl)prop-2-en-1-one (4g), purified on
silica gel column in methylene chloride, followed by crys-
tallization from ethanol, an orange solid, yield 41%, mp
144–145 °C.
(E)-1-(3-Oxido-5-propoxybenzo[d][1,3]oxathiol-6-yl)-3-
(2,4,6-trimethoxyphenyl)prop-2-en-1-one (4i), crystallization
solvent: methanol, a yellow solid, yield 45%, mp 183–
184 °C.
IR (KBr): 1651, 1607, 1563, 1460, 1329, 1206, 1124,
1060, 1006.
IR (KBr): 3230, 1641, 1555, 1523, 1440, 1269, 1134,
1011.
1H NMR (500 MHz, DMSO-d6): d 7.90 (S, 1H, H-4), 7.86
(d, 1H, J = 16.1 Hz, H-b), 7.41 (d, 1H, J = 16.1 Hz, H-a),
7.28 (s, 1H, H-7), 6.30 (s, 2H, H-30, H-50), 5.63 (d, 1H,
J = 11.4 Hz, H-2), 5.19 (d, 1H, J = 11.4 Hz, H-2), 4.06 (t,
1H NMR (500 MHz, DMSO-d6): d 9.26 (s, 1H, OH), 7.92
(s, 1H, H-4), 7.36 (m, 2H, H-7, H-b), 7.15 (m, 2H, H-20,
H-60), 7.08 (d, 1H, J = 15.9 Hz, H-a), 6.97 (d, 1H,
Chem Biol Drug Des 2016
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Konieczny et al.
2H, J = 6.3 Hz, OCH2), 3.86 (s, 9H, 3 9 OCH3), 1.68 (m,
2H, CH2), 0.90 (t, 3H, J = 7.4 Hz, CH3).
EA: Calcd for C23H26O7S: C, 61.87; H, 5.87; S, 7.18.
Found: C, 61.67; H, 5.81; S, 7.07.
13C NMR (126 MHz, DMSO-d6) d 193.69, 164.23, 162.04,
153.75, 152.41, 137.33, 136.08, 133.04, 125.91, 113.27,
112.00, 105.35, 91.97, 91.72, 71.33, 56.72, 56.28, 22.56,
10.95.
(E)-1-(5-Butoxy-3-oxidobenzo[d][1,3]oxathiol-6-yl)-3-(2,4,6-
trimethoxyphenyl)prop-2-en-1-one (4l), crystallization sol-
vent: methanol, a yellow solid, yield 74%, mp 180–181 °C.
IR (KBr): 1649, 1606, 1562, 1459, 1329, 1206, 1122,
1060, 1005.
EA: Calcd for C22H24O7S: C, 61.10; H, 5.59; S, 7.41.
Found: C, 60.86; H, 5.58; S, 7.45.
1H NMR (500 MHz, DMSO-d6): d 7.90 (s, 1H, H-4), 7.84
(d, 1H, J = 16.1 Hz, H-b), 7.40 (d, 1H, J = 16.1 Hz, H-a),
7.28 (s, 1H, H-7), 6.31 (s, 2H, H-30, H-50), 5.63 (d, 1H,
J = 11.4 Hz, H-2), 5.19 (d, 1H, J = 11.4 Hz, H-2), 4.09 (t,
1H, J = 6.3 Hz, OCH2), 3.86 (s, 9H, 3 9 OCH3), 1.63 (m,
2H, CH2), 1.36 (m, 2H, CH2), 0.81 (t, 3H, J = 7.4 Hz,
CH3).
(E)-3-(3-Fluoro-4-methoxyphenyl)-1-(3-oxido-5-propoxy-
benzo[d][1,3]oxathiol-6-yl)prop-2-en-1-one (4j), crystalliza-
tion solvent: ethanol, a cream solid, yield 84%, mp 158–
160 °C.
IR (KBr, cmꢀ1): 1662, 1608, 1592, 1516, 1283, 1130,
1041.
13C NMR (126 MHz, DMSO-d6) d 193.69, 164.21, 162.04,
153.73, 152.45, 137.28, 136.00, 133.05, 125.98, 113.27,
111.94, 105.36, 91.96, 91.68, 69.47, 56.72, 56.28, 31.18,
19.23, 14.23.
1H NMR (500 MHz, DMSO-d6): d 7.93 (s, 1H, H-4), 7.74
(dd, 1H, J1 = 12.7 Hz, J2 = 2.0 Hz, H-20), 7.56 (d, 1H,
J = 8.3 Hz, H-60), 7.43 (d, 1H, J = 16.1 Hz, H-b), 7.36 (s,
1H, H-7), 7.26 (d, 1H, J = 16.1 Hz, H-a), 7.22 (t, 1H,
J = 8.7 Hz, H-50), 5.65 (d, 1H, J = 11.2 Hz, H-2), 5.20 (d,
1H, J = 11.2 Hz, H-2),4.06 (t, 2H, J = 6.0 Hz, OCH2),
3.90 (s, 3H, OCH3), 1.67 (m, 2H, CH2), 0.88 (t, 3H,
J = 7.3 Hz, CH3).
EA: Calcd for C23H26O7S: C, 61.87; H, 5.87; S, 7.18.
Found: C, 61.63; H, 5.90; S, 6.99.
5-Acetyl-6-methoxy-3-oxidobenzo[d][1,3]oxathiole (9).
13C NMR (126 MHz, DMSO-d6) d 192.77, 153.69, 153.16,
152.62, 151.22, 150.04, 149.95, 143.92, 136.12, 133.44,
128.21, 128.15, 127.41, 127.39, 126.28, 115.86, 115.71,
114.54, 113.37, 112.05, 92.01, 71.28, 56.84, 22.61,
11.09.
About 30% Hydrogen peroxide (5 mL) was added to a
solution of 5-acetyl-6-methoxybenzo[d][1,3]oxathiole (8)
(18) (1.05 g, 5 mmol) in acetic acid (15 mL) and the mix-
ture was stirred at r.t. for 1 h. The solution was diluted
with water (100 mL) and extracted with methylene chloride
(3 9 30 mL). The organic layer was washed with water
(20 mL), dried (Na2SO4), evaporated to dryness, and the
residue was crystallized from methanol to give a colorless
solid, yield 46%, mp 157–158 °C.
EA: Calcd for C20H19FO5S: C, 61.53; H, 4.91; S, 8.21.
Found: C, 61.60; H, 4.86; S. 8.08.
(E)-1-(5-Butoxy-3-oxidobenzo[d][1,3]oxathiol-6-yl)-3-(3,4,5-
trimethoxyphenyl)prop-2-en-1-one (4k), crystallization sol-
vent: methanol, a yellow solid, yield 73%, mp 129–
130 °C.
IR (KBr): 1665, 1593, 1472, 1260, 1235, 1039.
1H NMR (500 MHz, DMSO-d6): d 8.29 (s, 1H, H-4), 7.13 (s,
1H, H-7), 5.67 (d, 1H, J = 5.4 Hz, H-2), 5.31 (d, 1H,
J = 5.4 Hz, H-2), 3.97 (s, 3H, OCH3), 2.53 (s, 3H, COCH3).
IR (KBr): 1655, 1578, 1504, 1464, 1419, 1274, 1128,
1010.
EA: Calcd for C10H10O4S: C, 53.09; H, 4.46; S, 14.17.
Found: C, 52.99; H, 4.40; S, 14.34.
1H NMR (500 MHz, DMSO-d6): d 7.95 (s, 1H, H-4), 7.42
(d, 1H, J = 16.0 Hz, H-b),7.36 (s, 1H, H-7), 7.34 (d, 1H,
J = 16.0 Hz, H-a), 7.11 (s, 2H, H-20, H-60), 5.65 (d, 1H,
J = 11.4 Hz, H-2), 5.20 (d, 1H, J = 11.4 Hz, H-2), 4.11 (t,
1H, J = 6.3 Hz, OCH2), 3.82 (s, 6H, 2 9 OCH3), 3.71 (s,
3H, OCH3), 1.65 (m, 2H, CH2), 1.36 (m, 2H, CH2), 0.79 (t,
3H, J = 7.4 Hz, CH3).
General procedure for condensation of 5-acetyl-6-
methoxy-3-oxidobenzo[d][1,3]oxathiole (9) with
benzaldehydes
A
mixture of 5-acetyl-6-methoxy-3-oxidobenzo[d][1,3]
oxathiole (9) (226 mg, 1 mmol), benzaldehyde
a
13C NMR (126 MHz, DMSO-d6) d 193.00, 153.75, 153.67,
152.66, 145.37, 140.43, 136.19, 133.37, 130.57, 126.81,
113.30, 112.02, 106.89, 92.01, 69.46, 60.81, 56.70,
31.28, 19.39, 14.21.
(1.2 mmol) and 5 N solution of NaOH (0.5 mL) in ethanol
(10 mL) was stirred in darkness for 3–4 h, the formed solid
was filtered off, washed with water and ethanol, and crys-
tallized.
4
Chem Biol Drug Des 2016

SAR of Cytotoxic Oxathiole-Fused Chalcones
(E)-1-(6-Methoxy-3-oxidobenzo[d][1,3]oxathiol-5-yl)-3-phe-
nylprop-2-en-1-one (5a), crystallization solvent: methanol,
a colorless solid, yield 41%, mp 155–157 °C.
EA: Calcd for C18H16O5S: C, 62.78; H, 4.68; S, 9.31.
Found: C, 62.74; H, 4.85; S, 9.12.
5-Acetoxy-3,3-dioxido-6-methoxybenzo[d][1,3]oxathiole
(11).
IR (KBr): 1651, 1603, 1573, 1469, 1245, 1068, 1037, 769.
1H NMR (500 MHz, DMSO-d6): d 8.27 (s, 1H, H-4), 7.76
(m, 2H, H-20, H-60), 7.57 (d, 1H, J = 16.1 Hz, H-b), 7.47
(m, 4H, H-a, H-30, H-40, H-50), 7.17 (s, 1H, H-7), 5.69 (d,
1H, J = 11.2 Hz, H-2), 5.33 (d, 1H, J = 11.2 Hz, H-2),
3.96 (s, 3H, OCH3).
About 30% Hydrogen peroxide (1 mL) was added to a
solution of 5-acetyl-6-methoxybenzo[d][1,3]oxathiole (8)
(18) (210 mg, 1 mmol) in acetic acid (3 mL) and the mix-
ture was stirred at 50 °C for 24 h. The precipitate was fil-
tered off, washed with water, dried, and crystallized from
2-methoxyethanol to give a colorless solid, yield 139 mg
(62%), mp 240–243 °C.
13C NMR (126 MHz, DMSO-d6) d 190.37, 164.62, 164.42,
143.48, 135.26, 131.26, 130.39, 129.70, 129.29, 127.15,
125.28, 123.65, 97.39, 93.37, 57.58.
IR (KBr): 1766, 1612, 1491, 1447, 1308, 1271, 1125, 910.
EA: Calcd for C17H14O4S: C, 64.95; H, 4.49; S, 10.20.
Found: C, 65.06; H, 4.45; S, 10.17.
1H NMR (500 MHz, DMSO-d6): d 7.71 (s, 1H, H-4), 7.10
(s, 1H, H-7), 5.42 (s, 2H, H-2), 3.86 (s, 3H, OCH3), 2.27
(s, 3H, OCOCH3).
(E)-3-(3-Chlorophenyl)-1-(6-methoxy-3-oxidobenzo[d][1,3]
oxathiol-5-yl)prop-2-en-1-one (5b), crystallization solvent:
methanol, a cream solid, yield 52%, mp 165–167 °C.
EA: Calcd C10H10O6S: Found: C, 46.48; H, 3.85; S,
12.35.
IR (KBr): 1645, 1599, 1568, 1465, 1418, 1304, 1246,
1064.
5-Acetyl-3,3-dioxido-6-methoxybenzo[d][1,3]oxathiole (12).
A mixture of 5-acetyl-6-methoxybenzo[d][1,3]oxathiole (8)
(630 mg, 3 mmol), potassium permanganate (1.92 g,
12 mmol), methylene chloride (18 mL) and water (36 mL)
was vigorously stirred at rt for 2 h. Next, the mixture was
decolorized by addition of aqueous sodium dithionite
(~1.5 g) solution, the organic layer was separated, and the
water layer was washed with methylene chloride
(2 9 30 mL). The combined organic layers were washed
with water (20 mL), dried (Na2SO4), and evaporated. Crys-
tallization of the residue from methanol gave a colorless
solid, yield 541 mg (74%), mp 205–208 °C.
1H NMR (500 MHz, DMSO-d6): d 8.29 (s, 1H, H-4), 7.88
(s, 1H, H-20), 7.74 (d, 1H, J = 7.3 Hz, H-60), 7.56 (d, 1H,
J = 16.8 Hz, H-b), 7.52 (d, 1H, J = 16.8 Hz, H-a), 7.48
(m, 2H, H-40, H-50), 7.17 (s, 1H, H-7), 5.69 (d, 1H,
J = 11.2 Hz, H-2), 5.33 (d, 1H, J = 11.2 Hz, H-2), 3.96 (s,
3H, OCH3).
13C NMR (126 MHz, DMSO-d6)
d 190.20, 164.73,
164.53, 141.62, 137.60, 134.46, 131.45, 130.75, 130.52,
128.82, 128.54, 127.81, 125.11, 123.64, 97.39, 93.40,
57.60.
IR (KBr): 1674, 1599, 1473, 1309, 1264, 1236, 1129,
1046.
EA: Calcd for C17H13ClO4S: C, 58.54; H, 3.76; S, 9.19.
Found: C, 58.16; H, 3.97; S, 8.96.
1H NMR (200 MHz, DMSO-d6): d 7.98 (s, 1H, H-4), 7.09
(s, 1H, H-7), 5.46 (s, 2H, H-2), 3.95 (s, 3H, OCH3), 2.51
(s, 3H, COCH3).
(E)-1-(6-Methoxy-3-oxidobenzo[d][1,3]oxathiol-5-yl)-3-(4-
methoxyphenyl)prop-2-en-1-one (5c), crystallization sol-
vent: ethanol, a cream solid, yield 53%, mp 180–182 °C.
EA: Calcd for C10H10O5S: C, 49.58; H, 4.16; S, 13.24.
Found: C, 49.53; H, 4.10; S, 13.44.
IR (KBr): 1650, 1597, 1571, 1510, 1255, 1174, 1066, 827.
1H NMR (500 MHz, DMSO-d6): d 8.22 (s, 1H, H-4), 7.71
(d, 2H, J = 8.8 Hz, H-20, H-60), 7.52 (d, 1H, J = 16.1 Hz,
H-b), 7.30 (d, 1H, J = 16.1 Hz, H-a), 7.15 (s, 1H, H-7),
7.00 (d, 2H, J = 8.8 Hz, H-30, H-50), 5.67 (d, 1H,
J = 11.7 Hz, H-2), 5.31 (d, 1H, J = 11.7 Hz, H-2), 3.94 (s,
3H, OCH3), 3.81 (s, 3H, OCH3).
5-Acetyl-3,3-dioxido-6-hydroxybenzo[d][1,3]oxathiole (14).
A mixture of 5-acetyl-6-hydroxybenzo[d][1,3]oxathiole (13)
(19) (294 mg, 1.5 mmol), potassium permanganate
(960 mg, 6 mmol), tetrabutylammonium bromide (120 mg,
0.4 mmol), methylene chloride (5 mL) and water (20 mL)
was vigorously stirred at rt for 1.5 h. Next, the mixture
was decolorized by addition of aqueous sodium dithionite
solution, the organic layer was separated, and the water
layer was washed with methylene chloride (2 9 20 mL).
The combined organic layers were washed with water
13C NMR (126 MHz, DMSO-d6)
d 190.40, 164.40,
164.23, 162.00, 143.72, 132.55, 131.18, 130.12, 127.82,
125.60, 124.82, 123.51, 115.18, 97.32, 93.33, 57.50,
56.06.
Chem Biol Drug Des 2016
5

Konieczny et al.
(20 mL), dried (Na2SO4), and evaporated. The residue
was purified on silica gel chromatographic column in
methylene chloride to give a colorless solid, yield 154 mg
(43%).
IR (KBr): 1676, 1463, 1321, 1170, 1141.
1H NMR (500 MHz, DMSO-d6): d 7.63 (s, 1H, H-4), 7.34
(s, 1H, H-7), 5.40 (s, 2H, H-2), 4.09 (t, 2H, J = 6.3 Hz,
OCH2), 2.54 (s, 3H, COCH3), 1.75 (m, 2H, CH2), 0.98 (t,
3H, J = 7.3 Hz, CH3).
IR (KBr): 3435, 1647, 1616, 1590, 1477, 1375, 1310,
1264, 1135, 1048.
EA: Calcd for C12H14O5S: C, 53.32; H, 5.22; S, 11.86.
Found: C, 53.20; H, 5.18; S, 11.51.
1H NMR (200 MHz, DMSO-d6): d 12.85 (bs, 1H, OH), 8.49
(s, 1H, H-4), 6.81 (s, 1H, H-7), 5.45 (s, 2H, H-2), 2.67 (s,
3H, COCH3).
General procedure for condensation of acetyl-3,3-
dioxidobenzo[d][1,3]oxathiole derivatives 12, 15,
18, and 19 with benzaldehydes
5-Acetyl-3,3-dioxido-6-(2-morpholinoethoxy)benzo[d][1,3]
oxathiole (15).
A mixture of suitable sulfone (1 mmol), a benzaldehyde
(1.2 mmol) and 5 N solution of NaOH or KOH (0.5 mL) in
ethanol (10 mL) was stirred in darkness for several hours
(3–20), the formed solid was filtered off, washed with
water and ethanol, and crystallized.
A mixture of 5-acetyl-3,3-dioxido-6-hydroxybenzo[d][1,3]
oxathiole (14) (360 mg, 1.58 mmol), 4-(2-chloroethyl)mor-
pholine hydrochloride (440 mg, 2.37 mmol), potassium
carbonate (830 mg, 6 mmol) in dry DMF (4 mL) was stir-
red at 60 °C for 4 h. The cooled reaction mixture was
diluted with water (100 mL) extracted with ethyl acetate
(3 9 30 mL). The organic layer was washed with water
(20 mL) and brine (20 mL), dried (MgSO4), and evapo-
rated. The residue was treated with ethyl ether, the formed
solid was filtered off and crystallized from methanol to give
a colorless solid, yield 46%.
(E)-1-(3,3-Dioxido-5-methoxybenzo[d][1,3]oxathiol-6-yl)-3-
phenylprop-2-en-1-one (6a), crystallization solvent: metha-
nol, a colorless solid, yield 66%, mp 161–162 °C.
IR (KBr): 1639, 1476, 1408, 1303, 1172, 1151, 1028.
1H NMR (500 MHz, DMSO-d6):
d
7.77 (dd, 2H,
IR (KBr): 1672, 1607, 1457, 1312, 1261, 1238, 1136,
1115, 1049.
J1 = 7.8 Hz, J2 = 2.0 Hz, H-20, H-60), 7.69 (s, 1H, H-4),
7.42–7.48 (m, 4H, H-30, H-40, H-50, H-b), 7.38 (s, 1H, H-
7), 7.25 (d, 1H, J = 16.1 Hz, H-a), 5.45 (s, 2H, H-2), 3.87
(s, 3H, OCH3).
1H NMR (200 MHz, DMSO-d6): d 7.97 (s, 1H, H-4), 7.14
(s, 1H, H-7), 5.47 (s, 2H, H-2), 4.30 (t, 2H, J = 5.4 Hz,
OCH2), 3.56 (m, 4H, O(CH2)2), 2.77 (t, 2H, J = 5.4 Hz,
NCH2), 2.58 (s, 3H, COCH3), 2.47 (m, 4H, N(CH2)2).
13C NMR (126 MHz, DMSO-d6) d 192.82, 153.63, 150.76,
146.18, 136.90, 134.86, 131.70, 129.71, 129.55, 127.07,
125.65, 115.87, 105.22, 83.77, 57.69.
EA: Calcd for C15H19NO6S: C, 52.77; H, 5.61; N, 4.10; S,
9.39. Found: 52.55; H, 5.50; N, 4.22; S, 9.19.
EA: Calcd for C17H14O5S: C, 61.81; H, 4.27; S, 9.71.
Found: C, 61.60; H, 4.45; S, 9.52.
6-Acetyl-3,3-dioxido-5-methoxybenzo[d][1,3]oxathiole (18).
(E)-3-(3-Chlorophenyl)-1-(3,3-dioxido-5-methoxybenzo[d]
[1,3]oxathiol-6-yl)prop-2-en-1-one (6b), crystallization sol-
vent: methanol, a cream solid, yield 82%, mp 170–
172 °C.
The reaction was done analogously to synthesis of com-
pound 14. The product was crystallized from methanol to
give a colorless solid, yield 64%, mp. 162–165 °C.
IR (KBr): 1680, 1476, 1404, 1312, 1169, 1142, 1030.
IR (KBr): 1644, 1597, 1475, 1410, 1308, 1150, 1028.
1H NMR (500 MHz, DMSO-d6): d 7.67 (s, 1H, H-4), 7.36
(s, 1H, H-7), 5.42 (s, 2H, H-2), 3.92 (s, 3H, OCH3), 2.53
(s, 3H, COCH3).
1H NMR (500 MHz, DMSO-d6): d 7.90 (s, 1H, H-20), 7.75
(d, 1H, J = 7.8 Hz, H-60), 7.69 (s, 1H, H-4), 7.44–7.52 (m,
3H, H-40, H-50, H-b), 7.38 (s, 1H, H-7), 7.33 (d, 1H,
J = 16.1 Hz, H-a), 5.44 (s, 2H, H-2), 3.87 (s, 3H, OCH3).
EA: Calcd for C10H10O5S: C, 49.58; H, 4.16; S, 13.24.
Found: C, 49.60; H, 4.05; S, 13.02.
EA: Calcd for C17H13ClO5S: C, 55.97; H, 3.59; S, 8.79.
Found: C, 55.81; H, 3.57; S, 8.53.
6-Acetyl-3,3-dioxido-5-propoxybenzo[d][1,3]oxathiole (19).
(E)-1-(3,3-Dioxido-5-methoxybenzo[d][1,3]oxathiol-6-yl)-3-
(4-methoxyphenyl)prop-2-en-1-one (6c) crystallization sol-
vent: methanol, a colorless solid, yield 95%, mp 197–
199 °C.
The reaction was done analogously to synthesis of com-
pound 29. The product was crystallized from ethanol to
give a colorless solid, yield 46%, mp. 170–172 °C.
6
Chem Biol Drug Des 2016

SAR of Cytotoxic Oxathiole-Fused Chalcones
IR (KBr): 1633, 1598, 1475, 1410, 1313, 1260, 1171,
1028.
(E)-1-(3,3-Dioxido-5-propoxybenzo[d][1,3]oxathiol-6-yl)-3-
(3-fluoro-4-methoxyphenyl)prop-2-en-1-one (6g), crystal-
lization from 2-methoxyethanol – ethanol mixture, a cream
solid, yield 57%, mp 178–179 °C.
1H NMR (500 MHz, DMSO-d6): d 7.72 (d, 2H, J = 8.8 Hz,
H-20, H-60), 7.66 (s, 1H, H-4), 7.40 (d, 1H, J = 16.1 Hz,
H-b), 7.34 (s, 1H, H-7), 7.09 (d, 1H, J = 16.1 Hz, H-a),
6.98 (d, 2H, J = 8.8 Hz, H-30, H-50), 5.44 (s, 2H, H-2),
3.86 (s, 3H, OCH3), 3.80 (s, 3H, OCH3).
IR (KBr, cmꢀ1): 1654, 1578, 1512, 1313, 1278, 1142,
1125, 1030.
1H NMR (500 MHz, DMSO-d6):
d
7.75 (dd, 1H,
EA: Calcd for C18H16O6S: C, 59.99; H, 4.47; S, 8.90.
Found: C, 59.83; H, 4.41; S, 8.90.
J1 = 12.7 Hz, J2 = 1.8 Hz, H-20), 7.67 (s, 1H, H-4), 7.57
(d, 1H, J = 8.6 Hz, H-60), 7.42 (d, 1H, J = 16.1 Hz, H-b),
7.35 (s, 1H, H-7), 7.22 (t, 1H, J = 8.6 Hz, H-50), 7.21 (d,
1H, J = 16.1 Hz, H-a), 5.44 (s, 2H, H-2), 4.08 (t, 2H,
J = 6.2 Hz, OCH2), 3.90 (s, 3H, OCH3), 1.64 (m, 2H,
CH2), 0.86 (t, 3H, J = 7.3 Hz, CH3).
(E)-1-(3,3-Dioxido-5-propoxybenzo[d][1,3]oxathiol-6-yl)-3-
(4-methoxyphenyl)prop-2-en-1-one (6d), crystallization sol-
vent: methanol, a cream solid, yield 46%, mp 148–
149 °C.
13C NMR (126 MHz, DMSO-d6) d 192.61, 153.15, 153.01,
151.20, 150.63, 150.12, 150.04, 144.71, 137.24, 128.14,
128.08, 127.51, 126.13, 125.65, 115.93, 115.86, 115.78,
114.53, 105.78, 83.75, 71.45, 56.86, 22.51, 11.03.
IR (KBr, cmꢀ1): 1665, 1599, 1570, 1513, 1423, 1312,
1155, 1032, 825.
1H NMR (500 MHz, DMSO-d6): d 7.72 (d, 2H, J = 8.6 Hz,
H-20, H-60), 7.66 (s, 1H, H-4), 7.44 (d, 1H, J = 16.0 Hz,
H-b), 7.34 (s, 1H, H-7), 7.15 (d, 1H, J = 16.0 Hz, H-a),
7.00 (d, 2H, J = 8.6 Hz, H-30, H-50), 5.44 (s, 2H, H-2),
4.08 (t, 2H, J = 6.2 Hz, OCH2), 3.82 (s, 3H, OCH3), 1.64
(m, 2H, CH2), 0.86 (t, 3H, J = 7.3 Hz, CH3).
EA: Calcd for C20H19FO6S: C, 59.10; H, 4.71; S, 7.89.
Found: C, 58.69; H, 4.60; S, 7.82.
(E)-1-(3,3-Dioxido-6-methoxybenzo[d][1,3]oxathiol-5-yl)-3-
phenylprop-2-en-1-one (7a), crystallization from methanol,
a cream solid, yield 66%, mp 180–183 °C.
13C NMR (126 MHz, DMSO-d6) d 192.54, 162.27, 152.97,
150.66, 145.83, 137.43, 131.38, 127.47, 125.56, 124.76,
115.85, 115.20, 105.75, 83.75, 71.44, 56.08, 22.51,
11.05.
IR (KBr, cmꢀ1): 1656, 1606, 1580, 1301, 1129, 1074.
1H NMR (500 MHz, DMSO-d6): d 8.00 (s, 1H, H-4), 7.76
(m, 2H, H-20, H-60), 7.55 (d, 1H, J = 15.6 Hz, H-b), 7.40–
7.47 (m, 4H, H-30, H-40, H-50, H-a), 7.14 (s, 1H, H-7), 5.49
(s, 2H, H-2), 3.95 (s, 3H, OCH3).
EA: Calcd for C20H20O6S: C, 61.84; H, 5.19; S, 8.25.
Found: C, 61.96; H, 5.23; S, 8.28.
(E)-1-(3,3-Dioxido-5-propoxybenzo[d][1,3]oxathiol-6-yl)-3-
(3,4,5-trimethoxyphenyl)prop-2-en-1-one (6f), crystallization
from 2-methoxyethanol – ethanol mixture, a cream solid,
yield 62%, mp 174–175 °C.
Calcd for C17H14O5S: C, 61.81; H, 4.27; S, 9.71. Found:
C, 61.90; H, 4.24; S, 9.51.
(E)-3-(4-Bromophenyl)-1-(3,3-dioxido-6-methoxybenzo[d]
[1,3]oxathiol-5-yl)prop-2-en-1-one (7b), crystallization from
2-methoxyethanol, a cream solid, yield 59%, mp 233–
234 °C.
IR (KBr, cmꢀ1): 1665, 1602, 1580, 1507, 1418, 1307,
1154, 1132, 1026.
1H NMR (500 MHz, DMSO-d6): d 7.68 (s, 1H, H-4), 7.41
(d, 1H, J = 16.1 Hz, H-b), 7.36 (s, 1H, H-7), 7.30 (d, 1H,
J = 16.1 Hz, H-a), 7.12 (s, 2H, H-20, H-60), 5.45 (s, 2H, H-
2), 4.09 (t, 2H, J = 6.2 Hz, OCH2), 3.82 (s, 6H,
2 9 OCH3), 3.71 (s, 3H, OCH3), 1.66 (m, 2H, CH2), 0.88
(t, 3H, J = 7.4 Hz, CH3).
IR (KBr, cmꢀ1): 1608, 1584, 1298, 1258, 1130, 1069.
1H NMR (500 MHz, DMSO-d6): d 8.01 (s, 1H, H-4), 7.74
(d, 2H, J = 8.5 Hz, H-20, H-60), 7.64 (d, 2H, J = 8.5 Hz,
H-30, H-50), 7.53 (d, 1H, J = 15.93 Hz, H-b), 7.45 (d, 1H,
J = 15.93 Hz, H-a), 7.14 (s, 1H, H-7), 5.49 (s, 2H, H-2),
3.94 (s, 3H, OCH3).
13C NMR (126 MHz, DMSO-d6) d 192.79, 153.76, 153.01,
150.62, 146.19, 140.52, 137.34, 130.46, 126.57, 125.59,
115.79, 106.99, 105.79, 83.76, 71.48, 60.81, 56.71,
22.54, 11.12.
13C NMR (126 MHz, DMSO-d6) d 190.18, 164.60, 161.56,
142.75, 134.47, 132.65, 131.29, 127.61, 126.01, 124.74,
124.26, 116.01, 99.56, 83.89, 57.82.
EA: Calcd for C22H24O8S: C, 58.92; H, 5.39; S, 7.15.
Found: C, 58.62; H, 5.41; S, 7.03.
Calcd for C17H13BrO5S: C, 49.89; H, 3.20; S, 7.84.
Found: C, 49.67; H, 3.28; S, 7.71.
Chem Biol Drug Des 2016
7

Konieczny et al.
(E)-3-(3-Chlorophenyl)-1-(3,3-dioxido-6-methoxybenzo[d]
[1,3]oxathiol-5-yl)prop-2-en-1-one (7c), crystallization from
methylene chloride – methanol mixture, a cream solid,
yield 74%, mp 192–195 °C.
(E)-3-(3-Fluoro-4-methoxyphenyl)-1-(6-(2-morpholi-
noethoxy)-3,3-dioxidobenzo[d][1,3]oxathiol-5-yl)prop-2-en-
1-one methanesulfonate hydrate (7f). Product of
condensation (free base) was obtained after crystallization
from chloroform
– methanol mixture, as an yellow
IR (KBr, cmꢀ1): 1657, 1608, 1579, 1476, 1311, 1257,
1131, 1072.
solid. The product (0.5 mmol) was dissolved in a solution
of methanesulfonic acid (1 mmol) in methylene chlo-
ride (2 mL). The formed clear solution was diluted
with ethyl ether, the precipitated solid was filtered off, and
washed with ethyl ether, to give a hygroscopic yellow
solid, yield 56%, mp 90 °C – softening, 145–150 °C
melting.
1H NMR (500 MHz, DMSO-d6): d 8.03 (s, 1H, H-4), 7.89
(s, 1H, H-20), 7.74 (d, 1H, H-60), 7.54 (d, 1H, J = 16.1 Hz,
H-b), 7.44–7.52 (m, 3H, H-40, H-50, H-a), 7.14 (s, 1H, H-
7), 5.49 (s, 2H, H-2), 3.94 (s, 3H, OCH3).
13C NMR (126 MHz, DMSO-d6) d 190.21, 164.64, 161.58,
142.38, 137.46, 134.47, 131.43, 130.88, 128.90, 128.30,
127.93, 125.98, 124.28, 115.99, 99.54, 83.89, 57.83.
IR (KBr, cmꢀ1): 3442, 1670, 1606, 1513, 1305, 1192,
1127, 1073.
1H NMR (500 MHz, DMSO-d6): d 9.91 (bs, 1H, NH+), 8.09
(s, 1H, H-4), 7.81 (d. 1H, J = 12.7 Hz, H-20), 7.59 (d, 1H,
J = 8.3 Hz, H-60), 7.50 (d, 1H, J = 16.1 Hz, H-b), 7.36 (d,
1H, J = 16.1 Hz, H-a), 7.23 (m, 2H, H-50, H-7), 5.51 (s,
2H, H-2), 4.56 (m, 2H, CH2), 3.90 (s, 3H, OCH3), 3.84 (m,
2H, CH2), 3.60 (m, 4H, 2 9 CH2), 3.45 (m, 2H, CH2), 3.12
(m, 2H, CH2).
EA: Calcd for C17H13ClO5S: C, 55.97; H, 3.59; S, 8.79.
Found: C, 55.67; H, 3.57; S, 8.93.
(E)-1-(3,3-Dioxido-6-methoxybenzo[d][1,3]oxathiol-5-yl)-3-
(4-methoxyphenyl)prop-2-en-1-one (7d), crystallization
from toluene, a cream solid, yield 54%, mp 210–212 °C.
IR (KBr, cmꢀ1): 1650, 1601, 1570, 1511, 1304, 1251,
1130, 1074, 827.
13C NMR (for a free base) (126 MHz, DMSO-d6) d 190.32,
163.66, 161.35, 153.19, 151.24, 149.92, 149.83, 142.95,
128.45, 128.40, 127.47, 126.44, 126.21, 124.07, 116.06,
115.68, 115.53, 114.53, 100.14, 83.86, 68.34, 66.70,
57.32, 56.88, 54.18.
1H NMR (500 MHz, DMSO-d6): d 7.96 (s, 1H, H-4), 7.72
(d, 2H, J = 8.8 Hz, H-20,H-60), 7.50 (d, 1H, J = 16.1 Hz,
H-b), 7.26 (d, 1H, J = 16.1 Hz, H-a), 7.13 (s, 1H, H-7),
7.00 (d, 2H, J = 8.8 Hz, H-30, H-50), 5.48 (s, 2H, H-2),
3.93 (s, 3H, OCH3), 3.81 (s, 3H, OCH3).
Calcd for C23H24FNO7S 9 CH3SO3H 9 H2O: Elemental
Analysis: Found: C, 48.37; H, 5.31; N, 2.09; S, 10.64.
Calcd for C18H16O6S: C, 59.99; H, 4.47; S, 8.90. Found:
C, 60.00; H, 4.46; S, 8.81.
(E)-1-(3,3-Dioxido-5-propoxybenzo[d][1,3]oxathiol-6-yl)-3-
(3-hydroxy-4-methoxyphenyl)prop-2-en-1-one (6e). A solu-
tion of Oxoneꢀ (20 g) in water (80 mL) was added dropwise,
during 2 h, to a stirred, boiling suspension of (E)-3-(3-
hydroxy-4-methoxyphenyl)-1-(5-propoxybenzo[d][1,3]
oxathiol-6-yl)prop-2-en-1-one (12) (3.72 g, 10 mmol) in
ethanol (400 mL). The obtained mixture was concentrated
under vacuum and diluted with water. The solid was filtered
off, dried and purified on silica gel column in methylene chlo-
ride – methanol 20:1 solution, and crystallized from ethanol
to give an yellow solid, yield 2.77 g (68%), mp 162–164 °C.
(E)-3-(3,4-Dimethoxyphenyl)-1-(3,3-dioxido-6-methoxy-
benzo[d][1,3]oxathiol-5-yl)prop-2-en-1-one (7e), crystalliza-
tion from 2-methoxyethanol, an yellow solid, yield 54%,
mp 218–220 °C.
IR (KBr, cmꢀ1): 1646, 1610, 1513, 1302, 1273, 1132,
1079, 1022.
1H NMR (500 MHz, DMSO-d6): d 7.94 (s, 1H, H-4), 7.45
(d, 1H, J = 16.0 Hz, H-b), 7.36 (s, 1H, H-20), 7.31 (d, 1H,
J = 8.3 Hz, H-60), 7.29 (d, 1H, J = 16.0 Hz, H-a), 7.13 (s,
1H, H-7), 7.01 (d, 1H, J = 8.3 Hz, H-50), 5.48 (s, 2H, H-2),
3.92 (s, 3H, OCH3), 3.81 (s, 6H, 2 9 OCH3).
IR (KBr, cmꢀ1): 3397, 1648, 1572, 1510, 1319, 1268,
1152, 1025.
1H NMR (500 MHz, DMSO-d6): d 9.25 (s, 1H, OH), 7.65
(s, 1H, H-4), 7.35 (m, 2H, H-b, H-7), 7.16 (m, 2H, H-20,
H-60), 7.03 (d, 1H, J = 16.0 Hz, H-a), 6.98 (d, 1H,
J = 8.2 Hz, H-50), 5.44 (s, 2H, H-2), 4.08 (t, 2H,
J = 5.8 Hz, OCH2), 3.83 (s, 3H, OCH3), 1.65 (m, 2H,
CH2), 0.87 (t, 3H, J = 7.6 Hz, CH3).
13C NMR (126 MHz, DMSO-d6) d 190.84, 164.24, 161.16,
151.99, 149.65, 145.27, 127.87, 126.65, 124.96, 124.04,
123.57, 115.82, 112.30, 111.59, 99.45, 83.86, 57.69,
56.31, 56.29.
Calcd for C19H18O7S: C, 58.45; H, 4.65; S, 8.21. Found:
C, 58.63; H, 4.55; S, 8.00.
13C NMR (126 MHz, DMSO-d6) d 192.29, 152.98, 151.28,
150.66, 147.40, 146.24, 137.40, 127.72, 125.60, 124.48,
8
Chem Biol Drug Des 2016

SAR of Cytotoxic Oxathiole-Fused Chalcones
122.92, 115.90, 114.95, 112.62, 105.77, 83.75, 71.46,
56.32, 22.53, 11.06.
USA). Cytotoxicity was determined upon comparison to
control cells.
EA: Calcd for C20H20O7S: C, 59.40; H, 4.98; S, 7.93.
Found: C, 59.12; H, 4.87; S, 7.68.
Tubulin polymerization assay
The test was performed according to manufacturer
instructions. Briefly, purified tubulin was diluted in the reac-
tion buffer (80 mM PIPES, pH 6.9, 2 mM MgCl2, 0.5 EGTA)
containing 1 mM GTP. Studied compounds were added at
respective concentrations and the kinetics of tubulin poly-
merization was determined by reading changes in 340 nm
absorbance with multiplate reader during 1 h at 37 °C.
Drug-response curves were normalized for buffer absor-
bance and drug concentrations inhibiting tubulin polymer-
ization by 50% were calculated using SLIDEWRITE (Advanced
Graphics Software, Inc., Rancho Santa Fe, CA, USA)
program.
Biology
Cells, drugs, and reagents
Human non-small lung adenocarcinoma A549 cells,
human colon carcinoma HCT-116 cells and human ovary
carcinoma HeLa cells used in these studies were obtained
from the American Type Culture Collection (ATCC; Rock-
ville, MD, USA). A549 cells were cultured in RPMI1640
medium, HCT-116 cells in McCoy’s 5 A medium and
HeLa cells in high glucose DMEM. All media were supple-
mented with 10% fetal bovine serum (FBS), antibiotics
(100 units/mL penicillin and 100 lg/mL streptomycin), and
cells were grown at 37 °C in a humidified 5% CO2-95%
air atmosphere.
Colchicine site competition assay
The assays were carried out using CytoDYNAMIX Screen
15 kit (Cytoskeleton Inc.) according to manufacturer’s
description. Briefly, biotinylated tubulin was bound to
streptavidin-coated yttrium SPA beads in G-PHEM buffer
for 30 min at 4 °C and exposed to 3H-colchicine in the
absence or presence of studied compounds. Radioactivity
of 3H-colchicine bound to tubulin was read using
MicroBeta2 scintillation counter (Perkin Elmer Wallac).
AMG compounds were dissolved in DMSO and kept at
ꢀ80 °C until use. Vinblastine, paclitaxel, colchicine were
from SIGMA-Aldrich-Fluka (Poznan, Poland). Radiolabeled
3H-colchicine, biotinylated colchicine as well as streptavidin-
coated yttrium beads were from Perkin Elmer. Tubulin Poly-
merization Kit and Colchicine Site Competitive Assay Kit
were from Cytoskeleton Inc. (Denver, CO, USA). Doxoru-
bicin was from the Institute of Biotechnology and Antibiotics
(Warsaw, Poland). All other reagents were of analytical
grade from either SIGMA-Aldrich-Fluka or POCH, S.A.
Results and Discussion
Chemistry
Mono-oxidized derivatives of chalcones (4) were prepared
by oxidation of related chalcones (1) (18) with hydrogen
peroxide in acetic acid at room temperature (Scheme 1).
The obtained products are listed in Table 1.
Cytotoxicity assays in vitro on A549, HCT116 and
HeLa cell lines
Multiwell (24–well) plates were seeded at 3 9 103 cells/
well in RPMI1640 medium supplemented with 10% FBS
and antibiotics (penicillin/streptomycin), and cells were
allowed to attach overnight. Drugs were added to wells in
10 lL aliquots of 200-times concentrated drug solutions
dissolved in DMSO in duplicates. Ten microliters of DMSO
was added to the control wells. Cells were incubated with
studied drugs for 72 h at 37 °C in a 5% CO2 atmosphere.
To all wells, 200 lL of MTT solution in PBS (4 mg/mL)
was added and incubated further for 3 h at 37 °C. Absor-
bance at 540 nm was measured after solubilization of for-
mazan crystals in 1 mL DMSO using a multiwell plate
reader (Victor3V; Perkin Elmer Wallac, Waltham, MA,
Isomeric sulfoxides 5 were prepared by oxidation of
acetophenone 8 (19) and condensation of the obtained
sulfoxide 9 with suitable benzaldehydes (Scheme 2). The
obtained products are listed in Table 2.
An attempt of preparation of the starting 6-acetyl-5-meth-
oxy-3,3-dioxidobenzo[d][1,3]oxathiole (12) by oxidation of
compound 10 (19) with hydrogen peroxide at elevated
temperature failed, as oxidation of the sulfur atom was
accompanied by Bayer – Villiger rearrangement to give
ester 11. The desired sulfone 12 was obtained by
O
R
O
R
O
X
S
X
S
30%H2O2
O
O
acetic acid, r.t.
O
O
4
1
Scheme 1: Synthesis of 3-oxidobenzo[d][1,3]oxathiol-6-yl)-3-phenylprop-2-en-1-ones (4).
Chem Biol Drug Des 2016
9

Konieczny et al.
Table 1: The obtained S-monoxido derivatives of chalcone (4) and their cytotoxic activity
O
R
O
X
S
O
O
Cytotoxicity, IC50 [lM] ꢁ SD
A549 HCT116
Influence on
activitya
Comp. No., Comp. code Substituent R Substituent X
HeLa
4a, AMG-225
4b, AMG-224
4c, AMG-221
4d, AMG-304
4e, AMG-302
4f, AMG-306
4g, AMG-332
4h, AMG-354
4i, AMG-335
4j, AMG-309
4k, AMG-339
4l, AMG-340
Doxorubicin
CA4
CH3
CH3
CH3
CH3
CH3
CH3
CH2CH2CH3
CH2CH2CH3
CH2CH2CH3
CH2CH2CH3
(CH2)3CH3
(CH2)3CH3
–
0.404 ꢁ 0.098 0.195 ꢁ 0.059
0.200 ꢁ 0.008 0.104 ꢁ 0.023
0.033 ꢁ 0.010 0.021 ꢁ 0.004
0.010 ꢁ 0.001 0.007 ꢁ 0.000
0.033 ꢁ 0.003 0.033 ꢁ 0.001
0.325 ꢁ 0.100 0.447 ꢁ 0.038
0.129 ꢁ 0.025
0.068 ꢁ 0.017
0.014 ꢁ 0.000
0.005 ꢁ 0.000
0.012 ꢁ 0.000
0.093 ꢁ 0.000
0.10
–
0.09
0.2
0.12
1.0
3-Cl
4-OCH3
3-OH-4-OCH3
3-F-4-OCH3
2,4,6-triOCH3
3-OH-4-OCH3
0.006 ꢁ 0.002 0.002 ꢁ 0.0005 0.002 ꢁ 0.0005 0.27
3-OPO3Na2-4-OCH3 0.010 ꢁ 0.003 0.003 ꢁ 0.001
0.003 ꢁ 0.001
0.038 ꢁ 0.011
0.005 ꢁ 0.000
2.25 ꢁ 0.258
0.018 ꢁ 0.004
0.154 ꢁ 0.017
0.003 ꢁ 0.000
0.008 ꢁ 0.003
0.08
0.03
0.06
0.19
0.02
2,4,6-triOCH3
3-F-4-OCH3
3,4,5-triOCH3
2,4,6-triOCH3
0.182 ꢁ 0.116 0.171 ꢁ 0.051
0.009 ꢁ 0.003 0.006 ꢁ 0.001
2.28 ꢁ 0.232
1.92 ꢁ 0.222
0.072 ꢁ 0.007 0.039 ꢁ 0.014
0.124 ꢁ 0.006 0.069 ꢁ 0.027
0.008 ꢁ 0.003 0.004 ꢁ 0.001
0.006 ꢁ 0.002 0.004 ꢁ 0.001
Pacilitaxel
SD, standard deviation.
aInfluence of oxidation on cytotoxic activity against A549 cells, calculated as a ratio of IC50 of oxidized analog 4 to IC50 of unoxidized form
1 (18).
X
OCH3
O
OCH3
OCH3
O
O
S
X
O
S
H
O C
O
S
30%H2O2
NaOH, ethanol, r.t.
acetic acid, r.t.
O
O
8
O
9
5
Scheme 2: Synthesis of 3-oxidobenzo[d][1,3]oxathiol-5-yl)-3-phenylprop-2-en-1-ones (5).
Table 2: The obtained S-monoxido derivatives of chalcone (5) and their cytotoxic activity
H
OC
X
3
O
S
O
O
Cytotoxicity, IC50 [lM] ꢁ SD
Influence on
activitya
Comp. No, Comp. code
Substituent X
A549
HCT116
HeLa
5a, AMG-212
5b, AMG-211
5c, AMG-186
–
4.81 ꢁ 0.56
1.56 ꢁ 0.09
6.39 ꢁ 0.06
2.40 ꢁ 0.57
0.960 ꢁ 0.040
4.90 ꢁ 0.23
4.29 ꢁ 0.69
1.63 ꢁ 0.14
3.57 ꢁ 0.06
1.2
0.8
18.1
3-Cl
4-OCH3
SD, standard deviation.
aInfluence of oxidation on cytotoxic activity against A549 cells, calculated as a ratio of IC50 of oxidized analog 5 to IC50 of unoxidized form 2 (19).
oxidation with potassium permanganate under phase-
transfer conditions (Scheme 3).
found that compound 14 could be obtained also by a
prolonged (2–3 days) oxidation with 30% H2O2 – acetic
acid at room temperature. Condensation of 14 with 4-
Similar oxidation of 5-acetyl-6-hydroxybenzo[d][1,3]oxathi-
ole (13) (19) gave the related dioxide 14. It was further
(2-chloroethyl)morpholine
(Scheme 4).
gave
compound
15
10
Chem Biol Drug Des 2016

SAR of Cytotoxic Oxathiole-Fused Chalcones
practically identical as for the parent compound. Oxidation
of the sulfoxides 4 to sulfones 6 also influenced signifi-
cantly the activity. For ring B unsubstituted derivatives
(compound 4a versus compound 6a) activity dropped by a
factor of 4, and for 4-OCH3 derivatives (compound 4c ver-
sus compound 6c) by a factor of 9. Yet, an opposite effect
was observed for 3-OH-4-OCH3 derivatives (compound
4g versus compound 6e) as oxidation to sulfone resulted
in twofold increase in activity. Generally, the tendency to
increase in activity prevailed, and three of the chalcones 6
H
OC
H
O
OC
3
O
S
3
O
S
30%H2O2
acetic acid, 50 oC
O
O
O
11
10
O
KMnO4
CH2Cl2 - H2O, r.t.
H
OC
3
O
S
O
O
12
O
demonstrated activity at nanomolar level (6d – IC50
=
0.004 lM; 6e – IC50 = 0.003 lM, and 6g – IC50 = 0.007
lM).
Scheme 3: Synthesis of 5-acetyl-3,3-dioxido-6-methoxybenzo[d]
[1,3]oxathiole (12).
Oxidation with potassium permanganate was utilized also
for preparation of sulfones 18 and 19 (Scheme 5).
For chalcones 2, 5, and 7, bearing the sulfur atom in posi-
tion meta to the carbonyl group, the influence of oxidation
on cytotoxic activity was not as clear as for the their iso-
mers 1, 4, and 6 discussed above. For ring B unsubsti-
tuted compounds 5a and 7a, oxidation almost did not
influence the activity. However, for 4-OCH3 derivatives 5c
and 7d, a sharp drop in activity was observed. Interest-
ingly, the effect was opposite to the one observed for iso-
mer with the sulfur atom para to the carbonyl group.
The described above sulfones 12, 15, 18 and 19 were
condensed with benzaldehydes under typical alkaline con-
ditions to give related chalcones (Scheme 6, Tables 3 and
4).
Surprisingly, the method failed in case of condensation
of 3-hydroxy-4-methoxybenzaldehyde with acetophenone
19. The reaction was relatively slow, and a prolonged
time was required to achieve an acceptable degree of
transformation of 19. As evidenced by TLC, the initially
formed chalcone 6e reacted further to give a mixture of
unidentified, colored products of low Rf, and the chal-
cone was isolated with yields below 10%. It could be
speculated that the side products were formed through
reactions of carbanions formed at carbon a to the sul-
fone group. For these reasons, the chalcone 6e was
prepared by Oxoneꢀ oxidation of related chalcone 1
(R = OCH2CH2CH3, X = 3-OH-4-OCH3).
Interaction with Tubulin
According to our early observations (19,23) the oxathiole-
fused chalcones, similarly as many other chalcones
(24–26) exhibit cytotoxic activity due to their influence on
polymerization of tubulin. It was of interest whether the
mechanism is also in operation in case of the oxidized
derivatives? To check it, inhibition of tubulin polymerization
and binding of the compounds at the colchicine site of
tubulin were studied experimentally (Table 5), and the
results proved that the oxidized compounds also interact
with tubulin.
Cytotoxic Activity In Vitro
Evaluation of cytotoxic activity of the prepared compounds
against three tumor cell lines using the MTT test
(Tables 1–4) demonstrated strong influence of oxidation on
activity (Tables 1–4, last columns). Oxidation of sulfur atom
in chalcones 1 to sulfoxides (chalcones 4) resulted in 5–10
times increase in activity, two of the compounds demon-
strated activity at nanomolar level (4g – IC50 = 0.006 lM;
4j – IC50 = 0.009 lM; all IC50 values used in this discus-
sion were measured in A549 cells). However, for the
2,4,6-trimethoxy derivative 4f, the IC50 value was
O
O
R
R
O
O
S
S
KMnO4
CH2Cl2 - H2O, r.t.
O
O
O
O
16 R = CH3
18 R = CH3
19 R = CH2CH2CH3
17 R = CH2CH2CH3
Scheme 5: Synthesis of 6-acetyl-5-alkoxy-3,3-dioxidobenzo[d]
[1,3]oxathioles 18 and 19.
N
O
H
O
H
O
O
O
O
S
O
S
O
S
ClCH2CH2NC4H8O
KMnO4
CH2Cl2 - H2O, r.t.
º
DMF, K2CO3, 70 C
O
O
O
O
O
O
15
14
13
Scheme 4: Synthesis of 5-acetyl-3,3-dioxido-6-(2-morpholinoethoxy)benzo[d][1,3]oxathiole (15).
Chem Biol Drug Des 2016
11

Konieczny et al.
X
R
O
R
O
X
NaOH orKOH
EtOH, rt
+
H
C O
O
O
12, 15, 18, 19
6a-g, 7a-f
Scheme 6: Synthesis of chalcones 6a–g and 7a–f.
Table 3: In vitro cytotoxicity of the prepared chalcones of general structure 6
O
O
R
O
X
S
O
O
Cytotoxicity, IC50 [lM] ꢁ SD
Influence on
activitya
Comp. No, Comp. code
Substituent R
Substituent X
A549
HCT116
HeLa
6a, AMG-229
6b, AMG-223
6c, AMG-228
6d, AMG-363
6e, AMG-358
6f, AMG-371
6g, AMG-370
CH3
CH3
CH3
CH2CH2CH3
CH2CH2CH3
CH2CH2CH3
CH2CH2CH3
–
3-Cl
4-OCH3
4-OCH3
3-OH-4-OCH3
3,4,5-triOCH3
30-F-40-OCH3
1.66 ꢁ 0.43
1.08 ꢁ 0.30
0.94 ꢁ 0.09
0.65 ꢁ 0.07
0.64 ꢁ 0.02
0.51 ꢁ 0.07
0.4
–
0.8
–
0.1
–
0.05
0.293 ꢁ 0.014
0.004 ꢁ 0.001
0.003 ꢁ 0.001
1.59 ꢁ 0.262
0.007 ꢁ 0.002
0.036 ꢁ 0.006
0.002 ꢁ 0.000
0.001 ꢁ 0.000
1.06 ꢁ 0.014
0.002 ꢁ 0.000
0.059 ꢁ 0.020
0.001 ꢁ 0.000
0.002 ꢁ 0.0005
1.55 ꢁ 0.071
0.001 ꢁ 0.000
SD, standard deviation.
aInfluence of oxidation on cytotoxic activity against A549 cells, calculated as a ratio of IC50 of oxidized analog 6 to IC50 of unoxidized form
1 (18).
Table 4: In vitro cytotoxicity of the prepared chalcones of general structure 7
R
X
O
O
S
O
O
O
Cytotoxicity, IC50 [lM] ꢁ SD
Influence
on activitya
Comp. No, Comp. code
Substituent R
Substituent X
A549
HCT116
HeLa
7a, AMG-213
7b, AMG-215
7c, AMG-214
7d, AMG-192
7e, AMG-216
7f, AMG-220ms
CH3
CH3
CH3
CH3
–
4-Br
3-Cl
4-OCH3
3,4-diOCH3
3-F-4-OCH3
3.05 ꢁ 0.42
1.60 ꢁ 0.14
1.07 ꢁ 0.31
1.87 ꢁ 0.14
9.59 ꢁ 1.99
6.86 ꢁ 0.27
2.34 ꢁ 0.36
1.43 ꢁ 0.18
0.88 ꢁ 0.20
0.91 ꢁ 0.11
11.64 ꢁ 2.76
4.27 ꢁ 0.17
2.83 ꢁ 0.13
1.60 ꢁ 0.01
1.10 ꢁ 0.13
0.48 ꢁ 0.02
7.24 ꢁ 0.80
6.19 ꢁ 1.08
0.7
0.5
0.5
5.3
8.0
1.6
CH3
CH2CH2morpholine
SD, standard deviation.
aInfluence of oxidation on cytotoxic activity against A549 cells, calculated as a ratio of IC50 of oxidized analog 7 to IC50 of unoxidized form
2 (19).
Our earlier molecular modeling evaluation of interaction of
selected chalcones 2 with tubulin indicated two preferen-
tial poses of the chalcones inside the colchicine binding
site (19). The first one, referred further as ‘A’ was adopted
by compounds 2 bearing alkoxy and hydroxy substituents
in the hydrophobic ring B. The second pose, referred fur-
ther as ‘B’, could be forced either by a presence of ami-
‘inappropriate’ substitution pattern of ring B, especially by
larger substituents, for example additional methoxy
groups, in positions 3 and 5, which disturb the interactions
which are characteristic for pose ‘A’. Prompted by the
results, we have performed a molecular modeling evalua-
tion of influence of oxidation of the sulfur atom on interac-
tions of the compounds with tubulin. As the detailed
procedure used to build complexes of chalcones with
noalkoxy group at position 20 of the ring
A or by
12
Chem Biol Drug Des 2016

SAR of Cytotoxic Oxathiole-Fused Chalcones
Table 5: Interaction of oxathiole-fused chalcones with tubulin
described in Ref. 19) is preferred. Since in the pose B, the
orientation of the benzoxathiole ring is flipped, oxidized
sulfur atom located in the para position (compounds 4 and
6) does not face the mentioned two beta strands, but is
placed in the proximity of the Cys241 residue what leads
to similar interactions like in the chalcones bearing the sul-
fur atom in meta position (compounds 5 and 7) described
below.
Inhibition of
polymerization
IC50 [lM]
Binding at
the colchicine
site IC50 [lM]
Comp. No,
Comp. label
Entry
1
2
3
4
5
6
7
8
4c
6c
4.55 ꢁ 0.4
7.35 ꢁ 0.3
2.80 ꢁ 0.4
1.96 ꢁ 0.3
17.38 ꢁ 0.7
4.15 ꢁ 0.3
2.23 ꢁ 0.3
1.73 ꢁ 0.2
0.195
0.164
0.021
0.191
4g
4j
4k
Such ligands (S-oxidized chalcones bearing the sulfur
atom in the position meta to the carbonyl group) exhibit
some differences in the interaction pattern with the binding
site with respect to their unoxidized counterparts. In this
case the oxidized ligand’s pose has to change slightly
since the additional oxygen replaces the sulfur in the inter-
action with the Cys241 thiol group (Figure 2 right). More-
over, nearby side chain of Leu248 residue becomes the
source of the steric hindrance. Thus, it seems that there
should be significant difference in activity of both possible
stereoisomers of respective sulfoxides. One with slightly
disrupted geometry but still able to form the hydrogen
bond with Cys241 and the other one deprived of this abil-
ity and heavily hindered by the Leu248 side chain. Unfor-
tunately, the above suggestion cannot be verified
experimentally since activities were measured for racemic
mixtures only. Further oxidation, leading to sulfones, may
surprisingly improve the binding slightly since, despite still
present unfavorable interaction of one of the oxygen atoms
with Leu248, the second oxygen atom can form favorable
interaction with Cys241, and counterbalance the unfavor-
able one with Leu248.
AMG-190a
Vinblastine
Combretastatin A4
0.108
0.023
aUnoxidized analog of compounds 4c and 6c of general formula
1; cytotoxic activity against A549: IC50 [lM] = 0.369 (19).
tubulin was already described (19), it was not repeated
here. As before, we have used flexible ligand docking with
the receptor held rigid, to assure comparability of the
results. It has to be noticed, that due to substantial flexibil-
ity of both loops and side chains within the colchicine
binding site, the method could result in some inaccuracy.
Based on the analysis of resulting complexes, S-oxidation
of chalcones bearing the sulfur atom in the position para
to the carbonyl group (compounds 4 and 6), and accept-
ing pose A in the colchicine binding site (as described in
Ref. 19), does not influence the mode of ligand binding. In
this ligand pose there is enough room around the chal-
cone’s sulfur atom, facing the beta strands 2 and 3 of the
beta subunit’s second domain, to accommodate the intro-
duction of even two additional oxygen atoms (Figure 2
left). Thus not only, there are no nearby groups causing
any steric hindrance, but additional oxygen atoms can
form favorable interactions with peptide bond dipoles of
both beta strands. All other important interactions, specific
for chalcones bound in the colchicine binding site in the
pose A, are preserved.
As demonstrated in Tables 1 and 3 (last columns) there
was a positive influence of oxidation of the sulfur atom
on cytotoxic activity. It was of interest if there was any
correlation among the following factors: cytotoxic activity
of the studied compounds – binding to tubulin at the col-
chicine site – inhibition of tubulin polymerization – and
results of docking to tubulin at the colchicine site?
However, that is not the case of derivatives with ring B
substituted in such a way, that binding in the pose B (as
Figure 2: Docking poses of ligands 6e (left) and 7d (right). Ligands are drawn as thick sticks while important residues in the binding site
as thinner sticks. Residue Val181 belongs to the a subunit of tubuline while all other shown residues belong to the subunit b.
Chem Biol Drug Des 2016
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Konieczny et al.
Comparison of data from Table 5 (entries 1, 2, and 6)
suggested that oxidation of the sulfur atom marginally
decreased binding to tubulin and inhibition of polymeriza-
tion while cytotoxic activity has changed in the opposite
direction, for the sulfoxide 4c even a ten times increase
in the activity was observed. The data are not contradic-
tory to conclusion from the molecular docking. Conse-
quently, it could be concluded that most probably the
observed influence of oxidation of the sulfur atom on
cytotoxic activity was not exclusively related to interaction
of the compounds with tubulin at the colchicine binding
site as molecular target. It remains to be established
whether the cytotoxic activity is influenced by an addi-
tional mechanism of the activity, differences in cell per-
meability, different metabolism, or still another cellular
effect.
4. Go M.L., Wu X., Liu X.L. (2005) Chalcones: an update
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A new group of oxathiole-fused chalcone derivatives active
at nanomolar level is described. It was found that oxidation
of sulfur atom of the oxathiole-fused chalcones strongly
influenced activity of the compounds, and that depending
on relative position of the sulfur atom in the molecule, the
activity could be either increased or diminished. Generally,
for the most active isomers with sulfur atom para to the
carbonyl group, oxidation led to increase of activity, while
for isomers with sulfur atom meta to the carbonyl the
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toxic activity of the compounds resulted, at least partly,
from their interaction with tubulin at the colchicine binding
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fur atom on cytotoxic activity cannot be solely related to
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Acknowledgments
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The Authors wish to thank the Polish Ministry of Science
and Higher Education for the grant no 0630/B/P01/2008/
34 (for M. Konieczny), and ADAMED GROUP for grants no
ONCO–3CLA/II–01/AMG (for M. Konieczny) and ONCO-
3CLA/CST/PG (for A. Skladanowski).
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Supporting Information
Additional Supporting Information may be found online in
the supporting information tab for this article:
Appendix S1. These data include H and 13C NMR spec-
1
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