Synthesis and antimitotic activity of alkoxy-substituted 1-aryl-3-(arylamino)alkenones

Article abstract of DOI:10.1007/s11172-015-0883-9

Alkoxy-substituted 1-aryl-3-(arylamino)prop-2-en-1-ones and 1-aryl-3-(arylamino)but-2en-1-ones were synthesized by the addition of anilines to 1,3-dioxo derivatives or aryl ethynyl ketones. The cis-trans isomerism of these compounds was studied. Biologica

Full text of DOI:10.1007/s11172-015-0883-9

Russian Chemical Bulletin, International Edition, Vol. 64, No. 2, pp. 439—444, February, 2015
439
Synthesis and antimitotic activity of alkoxyꢀsubstituted
1ꢀarylꢀ3ꢀ(arylamino)alkenones
A. V. Samet,a V. Yu. Zhuzhin,b M. N. Semenova,c and V. V. Semenova
aN. D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences,
47 Leninsky prosp., 119991 Moscow, Russian Federation.
Fax: +7 (495) 135 5328. Eꢀmail: sametav@server.ioc.ac.ru
bHigher Chemical College, Russian Academy of Sciences,
Build. 1, 9 Miusskaya pl., 125047 Moscow, Russian Federation
cN. K. Koltsov Institute of Developmental Biology, Russian Academy of Sciences,
26 ul. Vavilova, 119334 Moscow, Russian Federation
Alkoxyꢀsubstituted 1ꢀarylꢀ3ꢀ(arylamino)propꢀ2ꢀenꢀ1ꢀones and 1ꢀarylꢀ3ꢀ(arylamino)butꢀ2ꢀ
enꢀ1ꢀones were synthesized by the addition of anilines to 1,3ꢀdioxo derivatives or aryl ethynyl
ketones. The cisꢀtrans isomerism of these compounds was studied. Biological tests on a sea
urchin embryo model showed that 1ꢀarylꢀ3ꢀ(arylamino)propꢀ2ꢀenꢀ1ꢀones exhibit an antimitotic
effect both via microtubule destabilization and through the action on other cellular targets. The
antiproliferative activity of these compounds increases with an increase in the number of alkoxy
substituents in 1ꢀaryl ring, whereas the presence of one methoxy group in the para position
proved to be optimal for the arylamino moiety.
Key words: 1ꢀarylꢀ3ꢀ(arylamino)propꢀ2ꢀenꢀ1ꢀones, 1ꢀarylꢀ3ꢀ(arylamino)butꢀ2ꢀenꢀ1ꢀones,
cisꢀtrans isomerism, tubulin, antimitotic activity, sea urchin embryos.
The natural antimitotic agent combretastatin Aꢀ4
(CA4) is a powerful tubulin polymerization inhibitor that
binds at the colchicine site.1,2 The combretastatin derivaꢀ
tive phosphorylated at the hydroxy group is currently under
clinical trials as an antitumor drug.3,4 The simplicity of
the CA4 structure makes it possible to synthesize numerꢀ
ous analogs of this compound, which do not undergo
cisꢀtrans isomerism responsible for the loss of antimitotic
activity.5 In particular, earlier we have shown that such
compounds can be synthesized from allylpolyoxybenzenes
derived from plant raw materials (parsley and dill essenꢀ
tial oils).6—8
atom groups, such as —(CH2)2—, —CH2NH—, and
—CONH—,12—14 carbocycles or heterocycles (usually
fiveꢀmembered),15—18 oneꢀ and threeꢀatom moieties, such
as —C(=O)— and —C=C—C(=O)—,8,19—21 and some
other groups (structure I)9—11 can serve as the bridge X.
Fourꢀatom bridges in combretastatin analogs are less wellꢀ
studied. Notably, one of isomers of the CA4 analog
containing the butadiene moiety —C=C—C=C— in an
E,Z conformation proved to be a very active tubulin polyꢀ
merization inhibitor.22
Recently, the antitumor properties of CA4 analogs
containing another fourꢀatom group, viz., C(=O)—
CH=CH—NH, (structure II) have been investigated.23
Unexpectedly, it was found that the resulting (Z)ꢀ1ꢀarylꢀ
3ꢀarylaminopropꢀ2ꢀenꢀ1ꢀones II have the microtubuleꢀ
stabilizing rather than destabilizing effect, which is simiꢀ
lar to that exhibited by taxol, and is followed by caspaseꢀ
dependent apoptosis. In the cited study,23 the presence of
exclusively the Z isomer was proved by 1H NMR spectroꢀ
scopy in CDCl3; however, solutions for biological assays
were prepared in DMSO, which could lead to the isomerꢀ
ization typical of CA4 analogs.
Numerous studies9—11 demonstrated that the presence
of electronꢀdonating substituents OR or NR2 and the cis
arrangement of benzene rings with respect to the bridge X
that links these rings are key structural factors responsible
for the antimitotic activity of CA4 derivatives. Not only
the —C=C— bond that is present in CA4 but also twoꢀ
The aim of the present work was to synthesize 1ꢀarylꢀ
3ꢀ(arylamino)propꢀ2ꢀenꢀ1ꢀones 1 and 1ꢀarylꢀ3ꢀ(arylꢀ
amino)butꢀ2ꢀenꢀ1ꢀones 2, in which the benzene rings
are linked by the fourꢀatom —C(O)—CR=CH—NH—
fragment in order to investigate their stereoisomerꢀ
Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 2, pp. 0439—0444, February, 2015.
1066ꢀ5285/15/6402ꢀ0439 © 2015 Springer Science+Business Media, Inc.

440
Russ.Chem.Bull., Int.Ed., Vol. 64, No. 2, February, 2015
Samet et al.
10 ppm for the trans isomer. Meanwhile, only the cis isoꢀ
mer is observed in CDCl3. This is apparently due to the
stabilization of the cis isomer via an intramolecular
NꢀH...O=C hydrogen bond24—27 (this is confirmed also
by a downfield shift of the signal of the NH proton in the
spectrum of the cis isomer), which is partially cleaved in
the polar solvent DMSOꢀd6. Compounds 2 exist excluꢀ
sively as cis isomers.24 This follows from the chemical
shifts for the NH protons in the 1H NMR spectra of 2 (in
DMSOꢀd6), which are similar to those for the NH protons
in the cis isomers (but not in the trans isomers) of comꢀ
pounds 1. It should be noted that the isomeric composiꢀ
tion of products 1, which were prepared by the addition of
anilines to ethynyl ketones, may differ from that of the
same products prepared by the condensation of anilines
with benzoylacetaldehydes. For instance, according to
the 1H NMR spectroscopic data (in DMSOꢀd6, 1 h
after the dissolution), 1b generated from ethynyl ketone 4
(Ar = 4ꢀMeOC6H4) exists as a single isomer (cis). After
overnight storage, this isomer is transformed into a mixꢀ
ture of isomers of the same composition as for 1b, which
was prepared from acetophenone 3a through the correꢀ
sponding benzoylacetaldehyde (cis/trans is 7 : 3, this ratio
was observed both 1 and 24 h after the dissolution). Meanꢀ
I: X = CH2, CH2NH, CO, CONH, COCH=CH, CH=CH—CH=CH,
Het, etc.
R´ = H (1), Me (2)
ism and biological action as tubulin polymerization
modulators.
The target compounds were synthesized by the Claisen
condensation of the corresponding acetophenones 3 with
ethyl formate (ethyl acetate) to form benzoylacetaldehydes
(benzoylacetones) followed by the reaction of the latter
compounds with anilines. In some cases, related comꢀ
pounds were synthesized by the addition of anilines to aryl
ethynyl ketones 4 (Scheme 1).
1
while, according to the H NMR spectroscopic data (in
DMSOꢀd6), compound 1a exists as a mixture of isomers
of the cis/trans composition equal to 2 : 1 (5 min, 1 h, and
24 h after the dissolution) regardless of the method used
for the synthesis of 1a.
The biological activity of the compounds was examꢀ
ined on embryos of the sea urchin Paracentrotus lividus
using the phenotypic method developed in our earlier
study.28 This method allows for the detection of comꢀ
pounds capable of disturbing cell division and provides
information on the mechanism of antimitotic activity. The
characteristic change in the embryo motility — the spinꢀ
ning on the bottom of a vessel instead of forward swimꢀ
ming near the surface — attests to the ability of comꢀ
pounds to bind to tubulin and destabilize microtubules.28
1
The H NMR spectra recorded in DMSOꢀd6 (both 1
and 24 h after the dissolution) show that compounds 1 are
mixtures of cis and trans isomers (as a rule, with the former
predominating, except for compounds 1e,f containing subꢀ
stituents in the ortho position with respect to the bridging
C(O)—C=C—NH group), as evidenced by the spinꢀspin
coupling constants (12.4—13.0 Hz for the trans isomers
and 7.7—7.9 Hz for the cis isomers). The chemical shift of
the NH proton is ca. 12 ppm for the cis isomer and ca.
Scheme 1
R = H (1), Me (2)
1, 2
a
b
c
d
Ar
Ar´
3a—f, 4
Ar
4ꢀMeOC6H4
4ꢀMeOC6H4
4ꢀMeOC6H4
4ꢀMeOC6H4
3,4ꢀ(MeO)2C6H3
3,4,5ꢀ(MeO)3C6H2
2,5ꢀ(MeO)2ꢀ3,4ꢀ(OCH2O)C6H
2ꢀBrꢀ3,4ꢀ(MeO)2C6H2
4ꢀMeOC6H4
3,4,5ꢀ(MeO)3C6H2
4ꢀMeOC6H4
4ꢀMeOC6H4
4ꢀMeOC6H4
4ꢀMeOC6H4
a
b
c
d
e
f
3,4ꢀ(MeO)2C6H3
3,4,5ꢀ(MeO)3C6H2
2,5ꢀ(MeO)2ꢀ3,4ꢀ(OCH2O)C6H
2ꢀBrꢀ3,4ꢀ(MeO)2C6H2
4ꢀMeOC6H4
e
f
4

Antimitotic activity of arylaminopropenones
Russ.Chem.Bull., Int.Ed., Vol. 64, No. 2, February, 2015
441
Table 1. Action of alkoxyꢀsubstituted 1ꢀarylꢀ3ꢀarylaminoalkꢀ
enones on sea urchin embryos
firms the microtubuleꢀdestabilizing ability, was revealed
for the most active compound 1d. However, we failed to
observe embryo spinning in the case of compounds 1e and
1f, because the treatment with these compounds after
hatching caused embryo immobilization due to deciliaꢀ
tion. Compounds of series 2 exhibiting much lower antiꢀ
proliferative activity have a pronounced ability to cause
deciliation. It is known that antimitotic agents — tubulin
polymerization inhibitors (microtubuleꢀdestabilizing
agents) — do not suppress ciliary beating and cannot cause
the detachment of cilia from the cell surface.28—30 Thereꢀ
fore, compounds 1d, 1e, and 1f not only exhibit microꢀ
tubuleꢀdestabilizing activity but also act on other cellular
targets.
Comꢀ
pound
С*/mol L–1
Cleavage
Cleavage
alteration
Embryo Deciliaꢀ
arrest
spinning
tion
1b
2b
1d
2d
1е
2e
1f
СA4
2
2
0.02
4
0.05
4
0.2
0.002
>4
>4
0.2
>4
0.5
>4
2
>4
>4
2
>4
>4
>4
>4
0.05
>4
1
2
2
0.5
0.5
0.5
>4
0.01
It should be noted that our in vivo studies on sea urchin
embryos did not confirm the ability of compounds 1 to act
as promoters of tubulin polymerization and microtubuleꢀ
stabilizing agents, which has been found earlier in in vitro
experiments.23
It can be suggested that the configurational instability
of molecules 1 in solutions (cisꢀtrans isomerization) influꢀ
ences their biological activity, as it occurs in natural CA4.10
*C are the threshold concentrations causing the effect.
All the tested compounds exhibited the ability to supꢀ
press cell division, but they differed in the value and charꢀ
acter of the effect (Table 1). The activity of compounds of
series 1 increased with an increase in the number of methꢀ
oxy substituents in the ring A to three or four, whereas the
presence of one methoxy group in the para position proved
to be optimal for the ring B. Compounds 1d, 1e, and 1f
had a much stronger effect on the development of sea
urchin embryos compared to compound 1b and caused
not only the cleavage alteration but also the complete
cleavage arrest. In this case, sea urchin eggs acquired tuberꢀ
culate shape characteristic of microtubuleꢀdestabilizing
agents. The spinning of sea urchin embryos, which conꢀ
Experimental
1
The H NMR spectra of compounds 1 and 2 were recorded
in DMSOꢀd6 and CDCl3 on Bruker AM 300 and Bruker DRX
500 instruments operating at 300.13 and 500.13 MHz, respecꢀ
tively. The spectroscopic data are summarized in Table 2. The
yields of the products and the isomer ratios of compounds 1
Table 2. 1H NMR spectra of compounds 1a—f and 2a—e
Comꢀ
pound
1H NMR (, J/Hz)
C(2)H
C(3)H
NH
Other
1аa
6.02 (d,
J = 7.8, cis);
6.36 (d,
J = 12.4,
trans)
7.76 (dd,
J = 12.4,
J = 7.8, cis);
7.99 (br.s,
trans)
9.90 (br.s,
trans);
12.05 (d,
J = 12.4, cis)
3.73 (s, OMe, cis); 3.74 (s, OMe, trans); 3.82 (s, ОMе,
cis+trans); 6.93 (m, HAr, cis+trans); 7.02 (d, HAr, J = 8.5,
cis+trans); 7.10 (d, HAr, J = 8.6, trans); 7.26 (d, HAr, J = 8.7,
cis); 7.84 (d, HAr, J = 8.5, trans); 7.93 (d, HAr, J = 8.6, cis)
1аb
2aa
1ba
5.94 (d,
J = 7.8)
7.39 (dd,
J = 12.3,
J = 7.8)
12.11 (d,
J = 12.3)
3.80 (s, 3 H, OMe); 3.87 (s, 3 H, ОMе); 6.89 (d, 2 H, HAr,
J = 8.9); 6.94 (d, 2 H, HAr, J = 8.8); 7.04 (d, 2 H, HAr, J = 8.9);
7.92 (d, 2 H, HAr, J = 8.8)
5.97 (s)
—
12.90 (s)
2.08 (s, 3 H, Me); 3.77 (s, 3 H, OMe); 3.82 (s, 3 H, OMe);
6.98 (m, 4 Н, НAr); 7.21 (d, 2 H, HAr, J = 8.7);
7.89 (d, 2 H, HAr, J = 8.8)
6.06 (d,
J = 7.9, cis);
6.41 (d,
J = 12.5,
trans)
7.88 (dd,
J = 12.3,
J = 7.9, cis);
8.08 (br.s,
trans)
9.93 (br.s,
trans);
11.98 (d,
J = 12.4, cis)
3.61 (s, OMe, trans); 3.62 (s, OMe, cis);
3.80—3.84 (m, ОMе, cis+trans); 6.42 (s, HAr, trans);
6.63 (s, HAr, cis); 7.03 (m, HAr, cis+trans);
7.85 (d, HAr, J = 8.8, trans); 7.95 (d, HAr, J = 8.8, cis)
1ba,c
6.06 (d,
J = 7.9)
7.88 (dd,
J = 12.3,
J = 7.9)
11.98 (d,
J = 12.4)
3.62 (s, 3 H, OMe); 3.82 (s, 6 H, OMe);
3.83 (s, 3 H, OMe); 6.63 (s, 2 H, HAr);
7.02 (d, 2 H, HAr, J = 8.8); 7.95 (d, 2 H, HAr, J = 8.8)
(to be contined)

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Russ.Chem.Bull., Int.Ed., Vol. 64, No. 2, February, 2015
Samet et al.
Table 2 (continued)
Comꢀ
1H NMR (, J/Hz)
pound
C(2)H
C(3)H
NH
Other
1bb
2ba
1ca
5.98 (d,
J = 7.9)
7.41 (dd,
J = 12.2,
J = 7.9)
12.12 (d,
J = 12.2)
3.82 (s, 3 H, OMe); 3.87 (s, 9 H, OMe);
6.31 (s, 2 H, HAr); 6.95 (d, 2 H, HAr, J = 8.8);
7.93 (d, 2 H, HAr, J = 8.8)
6.02 (s)
—
12.98 (s)
2.19 (s, 3 H, Me); 3.66 (s, 3 H, OMe); 3.80 (s, 6 H, OMe);
3.82 (s, 3 H, OMe); 6.58 (s, 2 Н, НAr);
7.00 (d, 2 H, HAr, J = 8.8); 7.91 (d, 2 H, HAr, J = 8.8)
6.06 (d,
J = 7.8,
cis);
6.38 (d,
J = 12.4,
trans)
7.76 (dd,
J = 12.4,
J = 7.8, cis);
8.00 (br.s,
trans)
9.88 (br.s,
trans);
12.08 (d,
J = 12.5, cis)
3.73 (s, OMe, trans); 3.75 (s, OMe, cis);
3.81—3.83 (m, ОMе, cis+trans); 6.91—6.96 (m, HAr,
cis+trans); 7.02—7.05 (m, HAr, cis+trans);
7.25 (d, HAr, J = 9.0, cis); 7.43 (d, HAr, J = 2.0, trans);
7.47 (dd, HAr, J = 8.4, J = 2.0, trans); 7.50 (d, HAr,
J = 2.0, cis); 7.58 (dd, HAr, J = 8.4, J = 2.0, cis)
1cb
2ca
5.95 (d,
J = 7.8)
7.40 (dd,
J = 12.4,
J = 7.8)
12.12 (d,
J = 12.4)
3.80 (s, 3 H, OMe); 3.94 (s, 3 H, OMe); 3.96 (s, 3 H,
ОMе); 6.89 (m, 3 H, HAr); 7.54 (dd, 1 H, HAr,
J = 8.4, J = 2.0); 7.56 (d, 1 H, HAr, J = 2.0)
6.01 (s)
—
12.92 (s)
2.11 (s, 3 H, Me); 3.78 (s, 3 H, OMe);
3.82 (s, 6 H, OMe); 6.97 (d, 2 H, HAr, J = 8.8);
7.01 (d, 1 H, HAr, J = 8.4); 7.21 (d, 2 H, HAr, J = 8.8);
7.48 (s, 1 H, HAr); 7.54 (d, 2 H, HAr, J = 8.4)
1da
6.11 (d,
J = 7.8, cis);
6.37 (d,
7.80 (dd,
J = 12.5,
J = 7.8, cis);
9.99 (br.s,
trans);
12.15 (d,
3.72—3.75 (m, OMe, cis+trans); 3.85—3.86 (m, OMe,
cis+trans); 6.92 (d, HAr, J = 9.0, trans); 6.95 (d, HAr, J = 9.0,
cis); 7.11 (d, HAr, J = 9.0, trans); 7.15 (s, HAr, trans);
7.24 (s, HAr, cis); 7.28 (d, HAr, J = 9.0, cis)
J = 12.4, trans) 8.03 (br.s, trans) J = 12.6, cis)
1db
2da
1ea
5.93 (d,
J = 7.7)
7.44 (dd,
J = 12.4,
J = 7.7)
12.17 (d,
J = 12.4)
3.81 (s, 3 H, OMe); 3.91 (s, 3 H, OMe);
3.94 (s, 6 H, OMe); 6.90 (d, 2 H, HAr, J = 8.9);
7.05 (d, 2 H, HAr, J = 8.9); 7.20 (s, 2 H, HAr);
6.09 (s)
—
13.0 (s)
2.11 (s, 3 H, Me); 3.72 (s, 3 H, OMe); 3.78 (s, 3 H, OMe);
3.85 (s, 6 H, OMe); 6.98 (d, 2 H, HAr, J = 8.9);
7.21 (s, 2 H, HAr); 7.23 (d, 2H, HAr, J = 8.9)
5.97 (d,
J = 7.8, cis);
6.16 (d,
J = 12.5,
trans)
7.74 (dd,
J = 12.5,
J = 7.8, cis);
7.85 (br.s,
trans)
9.88 (br.s,
trans);
11.96 (d,
J = 12.5, cis)
3.72 (s, OMe, trans); 3.74 (s, OMe, cis); 3.80 (s, OMe, trans);
3.81 (s, OMe, cis); 3.82 (s, OMe, trans), 3.86 (s, OMe, cis);
6.09 (s, SН2, trans); 6.11 (s, SН2, cis); 6.80 (s, HAr, trans);
6.91 (d, HAr, J = 8.9, trans); 6.94 (d, HAr, J = 8.9, cis);
6.98 (s, HAr, cis); 7.05 (d, HAr, J = 8.9, trans);
7.26 (d, J = 8.9, HAr, cis)
1eb
2ea
6.09 (d,
J = 7.7)
7.36 (dd,
J = 12.5,
J = 7.7)
12.06 (d,
J = 12.5)
3.80 (s, 3 H, OMe); 3.91 (s, 3 H, OMe); 3.94 (s, 3 H, OMe);
6.05 (s, 2 H, SH2); 6.89 (d, 2 H, HAr, J = 8.9);
7.04 (d, 2 H, HAr, J = 8.9); 7.07 (s, 1 H, HAr)
5.82 (s)
—
12.80 (s)
2.02 (s, 3 H, Me); 3.78 (s, 3 H, OMe); 3.80 (s, 3 H, OMe);
3.85 (s, 3 H, OMe); 6.09 (s, 2 H, SH2);
6.91 (s, 1 H, HAr); 6.97 (d, 2 H, HAr, J = 8.9);
7.22 (d, 2 H, HAr, J = 8.9)
1fa
5.64 (d,
J = 7.7, cis);
5.78 (d,
J = 13.0,
trans)
7.79 (dd,
J = 12.7,
J = 7.7, cis);
7.60 (br.s,
trans)
10.0 (br.s,
trans);
11.81 (d,
J = 12.7, cis)
3.71 (s, OMe, trans); 3.75 (s, OMe, cis); 3.77 (s, OMe, trans);
3.79 (s, OMe, cis); 3.81 (s, OMe, trans); 3.82 (s, OMe, cis);
6.90 (d, HAr, J = 8.7, trans); 6.94—7.03 (m, HAr,
cis+trans); 7.08 (s, HAr, cis); 7.16 (d, HAr, J = 8.9, trans);
7.29 (d, HAr, J = 8.9, cis)
1fb
5.74 (d,
J = 7.6)
7.36 (dd,
J = 12.6,
J = 7.6)
12.06 (d,
J = 12.6)
3.81 (s, 3 H, OMe); 3.90 (s, 3 H, OMe); 3.91 (s, 3 H, OMe);
6.90 (d, 2 H, HAr, J = 8.9); 7.05 (s, 1 H, HAr);
7.06 (d, 2 H, HAr, J = 8.9); 7.08 (s, 1 H, HAr)
a The spectrum was recorded in DMSOꢀd6.
b The spectrum was recorded in CDCl3.
c The product was obtained from ketone 4; the spectrum was recorded one hour after the dissolution.

Antimitotic activity of arylaminopropenones
Russ.Chem.Bull., Int.Ed., Vol. 64, No. 2, February, 2015
443
(according to the 1H NMR spectroscopic data in DMSOꢀd6) are
given below.
1ꢀ(3,4,5ꢀTrimethoxyphenyl)ꢀ3ꢀ(4ꢀmethoxyphenylamino)propꢀ
2ꢀenꢀ1ꢀone (1d). Yield 72%, cis/trans ratio is 2 : 1. M.p.
101—103 C (cf. lit. data23: m.p. 107—108 C). Found (%):
C, 66.75; H, 5.91; N, 3.85. C19H21NO5. Calculated (%):
C, 66.46; H, 6.16; N, 4.08.
1ꢀ(3,4,5ꢀTrimethoxyphenyl)ꢀ3ꢀ(4ꢀmethoxyphenylamino)butꢀ
2ꢀenꢀ1ꢀone (2d). Yield 59%. M.p. 112 C. Found (%): C, 66.93;
H, 6.57; N, 4.14. C20H23NO5. Calculated (%): C, 67.21; H,
6.49; N, 3.92.
1ꢀ[2,5ꢀDimethoxyꢀ3,4ꢀ(methylenedioxy)phenyl]ꢀ3ꢀ(4ꢀmethꢀ
oxyphenylamino)propꢀ2ꢀenꢀ1ꢀone (1e). Yield 60%, cis/trans ratio
is 6 : 5. M.p. 118—119 C. Found (%): C, 64.13; H, 5.17; N, 4.18.
C19H19NO6. Calculated (%): C, 63.86; H, 5.36; N, 3.92.
1ꢀ[2,5ꢀDimethoxyꢀ3,4ꢀ(methylenedioxy)phenyl]ꢀ3ꢀ(4ꢀmethꢀ
oxyphenylamino)butꢀ2ꢀenꢀ1ꢀone (2e). Yield 43%. M.p. 93—95 C.
Found (%): C, 64.85; H, 5.52; N, 3.89. C20H21NO6. Calculatꢀ
ed (%): C, 64.68; H, 5.70; N, 3.77.
1ꢀ(2ꢀBromoꢀ4,5ꢀdimethoxyphenyl)ꢀ3ꢀ(4ꢀmethoxyphenylꢀ
amino)propꢀ2ꢀenꢀ1ꢀone (1f). Yield 58%, cis/trans ratio is 2 : 3.
M.p. 130—132 C. Found (%): C, 54.91; H, 4.66; Br, 20.08;
N, 3.77. C18H18BrNO4. Calculated (%): C, 55.12; H, 4.63;
Br, 20.37; N, 3.57.
Investigation of the antiproliferative activity of compounds on
the sea urchin embryo model.28 Experiments were carried out in
the Biological laboratory of the N. K. Koltsov Institute of Deꢀ
velopmental Biology of the Russian Academy of Sciences in
Cyprus. Adult sea urchins Paracentrotus lividus L. (Echinidae,
Echinodermata) were collected in the coastal zone and kept in an
aerated seawater aquarium. The spawning was induced by inꢀ
jecting sea urchins with 0.5 M KCl (1—2 mL). The eggs were
washed with seawater filtered through a nylon filter and fertilꢀ
ized by the addition of several drops of dilute sperm. Embryos
(600—2000 per milliliter) were incubated in filtered seawater at
room temperature (18—23 C) to the stage of beginning of active
feeding (36—40 h, middle pluteus 2). Throughout the incubation
period, the embryo suspension was stirred using a plastic paddle
at a rotation speed of 60 rpm driven by an electric motor.
The stock solutions of chemical compounds were prepared
in DMSO and 96% ethanol; the maximum studied concentraꢀ
tions of the compounds depended on their solubility. The soluꢀ
bility of the compounds in solvents and seawater was monitored
with a microscope.
The compounds were tested in 6ꢀwell cell culture plates at
the following stages of the development: 1) eggs (10—15 min
after the fertilization); 2) freeꢀswimming hatched blastula
(9—10 h after the fertilization). An egg suspension (5 mL) was
placed in each well, and the corresponding volume of the soluꢀ
tion of the compound tested was added to the required final
concentration. The maximum concentration of the solvent did
not exceed the maximal tolerated concentration (1% for ethanol
and 0.05% for DMSO). The tests were done by twofold serial
dilutions of the compounds until the effect disappeared. The
antiproliferative activity of the compounds was estimated as the
lowest (threshold) concentration that alters fertilized egg diviꢀ
sion. A change in the motility of sea urchin embryos was evaluatꢀ
ed after the treatment with the compounds under study at
the stage of hatched blastula, when embryos started to swim
due to coordinated beating of cilia. The suppression of forward
swimming, the settlement to the bottom of the vessel, and
the rapid spinning of embryos attest to the ability of the comꢀ
pound under study to destabilize microtubules. The observaꢀ
Synthesis of 1ꢀarylꢀ3ꢀarylaminopropꢀ2ꢀenꢀ1ꢀones 1a,b from
4ꢀmethoxyphenyl ethynyl ketone (4) (general procedure). A soluꢀ
tion of compound 4 (1 mol) and the corresponding aniline (1 mol)
in ethanol was refluxed for 5 min and then cooled. The bright
yellow crystalline precipitate that formed was filtered off.
Synthesis of 1ꢀarylꢀ3ꢀarylaminopropꢀ2ꢀenꢀ1ꢀones 1a—f (genꢀ
eral procedure). Sodium hydride (60% suspension in mineral oil,
80 mg, 2 mmol) was added to a solution of the corresponding
acetophenone 1 (2 mmol) and HCOOEt (2 mmol) in THF
(3 mL). The reaction mixture was stirred for 4 h (in the case of
acetophenones 3e,f, for 18 h) and diluted with diethyl ether
(3 mL). The resulting suspension was filtered off, and the preꢀ
cipitate was washed with diethyl ether (5 mL) and dried. The
Na salt of the corresponding aroylacetaldehyde thus formed was
dissolved in EtOH (3 mL). Then the corresponding aniline
(2 mmol) and several drops of AcOH were added. The resulting
solution was refluxed for 5 min and then cooled. The bright
yellow crystalline precipitate that formed was filtered off.
Synthesis of 1ꢀarylꢀ3ꢀarylaminobutꢀ2ꢀenꢀ1ꢀones 2a—e (genꢀ
eral procedure). Sodium hydride (60% suspension in mineral oil,
80 mg, 2 mmol) was added to a solution of the corresponding
acetophenone 1 (2 mmol) and MeCOOEt (2 mmol) in THF
(3 mL). The reaction mixture was stirred for 8 h (in the case of
acetophenone 3e, for 24 h) and diluted with diethyl ether (3 mL).
The resulting suspension was filtered off, and the precipitate was
washed with diethyl ether (5 mL) and dried. The Na salt of the
corresponding aroylacetone thus formed was dissolved in EtOH
(3 mL). Then the corresponding aniline (2 mmol) and several
drops of AcOH were added. The resulting solution was refluxed
for 3 h and then cooled. The pale yellow crystalline precipitate
that formed was filtered off. Product 2e was initially obtained as
an oil, which was purified by flash chromatography on silica gel
(hexane—ethyl acetate, 5 : 1, as the eluent).
1ꢀ(4ꢀMethoxyphenyl)ꢀ3ꢀ(4ꢀmethoxyphenylamino)propꢀ2ꢀenꢀ
1ꢀone (1a). Yield 88% (from ethynyl ketone 4), 69% (from aceꢀ
tophenone 3a); cis/trans ratio is 2 : 1. M.p. 185—187 C (cf. lit.
data23: m.p. 187—188 C). Found (%): C, 71.83; H, 5.90; N, 5.18.
C17H17NO3. Calculated (%): C, 72.07; H, 6.05; N, 4.94.
1ꢀ(4ꢀMethoxyphenyl)ꢀ3ꢀ(4ꢀmethoxyphenylamino)butꢀ2ꢀenꢀ
1ꢀone (2a). Yield 61%. M.p. 144—145 C (cf. lit. data31: m.p.
142—143 C). Found (%): C, 72.96; H, 6.35; N, 4.88.
C18H19NO3. Calculated (%): C, 72.71; H, 6.44; N, 4.71.
1ꢀ(4ꢀMethoxyphenyl)ꢀ3ꢀ(3,4,5ꢀtrimethoxyphenylamino)propꢀ
2ꢀenꢀ1ꢀone (1b). Yield 82% (from ethynyl ketone 4, cis isomer),
65% (from acetophenone 3a, cis/trans ratio is 7 : 3). M.p.
123—124 C. Found (%): C, 66.09; H, 5.94; N, 4.29. C19H21NO5.
Calculated (%): C, 66.46; H, 6.16; N, 4.08.
1ꢀ(4ꢀMethoxyphenyl)ꢀ3ꢀ(3,4,5ꢀtrimethoxyphenylamino)butꢀ
2ꢀenꢀ1ꢀone (2b). Yield 55%. M.p. 103—105 C. Found (%):
C, 66.89; H, 6.58; N, 4.13. C20H23NO5. Calculated (%):
C, 67.21; H, 6.49; N, 3.92.
1ꢀ(3,4ꢀDimethoxyphenyl)ꢀ3ꢀ(4ꢀmethoxyphenylamino)propꢀ2ꢀ
enꢀ1ꢀone (1c). Yield 70%, cis/trans ratio is 2 : 1. M.p. 155—156 C.
Found (%): C, 69.24; H, 5.95; N, 4.41. C18H19NO4. Calculatꢀ
ed (%): C, 68.99; H, 6.11; N, 4.47.
1ꢀ(3,4ꢀDimethoxyphenyl)ꢀ3ꢀ(4ꢀmethoxyphenylamino)butꢀ2ꢀ
enꢀ1ꢀone (2c). Yield 62%. M.p. 135—136 C. Found (%):
C, 70.00; H, 6.30; N, 4.04. C19H21NO4. Calculated (%):
C, 69.71; H, 6.47; N, 4.28.

444
Russ.Chem.Bull., Int.Ed., Vol. 64, No. 2, February, 2015
Samet et al.
tions were made with a Biolam LOMO optical microscope
(StꢀPetersburg).
16. L. Wang, K. W. Woods, Q. Li, K. J. Barr, R. W. McCroskey,
S. M. Hannick, L. Gherke, R. B. Credo, Y.ꢀH. Hui,
K. Marsh, R. Warner, J. Y. Lee, N. ZielinskiꢀMozng,
D. Frost, S. H. Rosenberg, H. L. Sham, J. Med. Chem.,
2002, 45, 1697.
This study was financially supported by the Russian
Foundation for Basic Research (Project No. 13ꢀ03ꢀ90455).
17. D. V. Tsyganov, L. D. Konyushkin, I. B. Karmanova, S. I.
Firgang, Yu. A. Strelenko, M. N. Semenova, A. S. Kiselyov,
V. V. Semenov, J. Nat. Prod., 2013, 76, 1485.
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Received July 29, 2014