N-(4-Methoxyphenyl)-substituted bicyclic isothioureas: effect on morphology of cancer cells

Article abstract of DOI:10.1016/j.mencom.2021.04.003

Bicyclic isothioureas of N-(4-methoxyphenyl)-2-aminocycloalkane[d]thiazole type were obtained using intramolecular electrophilic cyclization of N-(cycloalk-2-enyl)-N′-(4-methoxyphenyl)thioureas. Isothiourea fused with sevenmembered ring caused noticeable

Full text of DOI:10.1016/j.mencom.2021.04.003

Mendeleev
Communications
Mendeleev Commun., 2021, 31, 288–290
N-(4-Methoxyphenyl)-substituted bicyclic isothioureas:
effect on morphology of cancer cells
Anna V. Evdokimova,a Alexander A. Alexeev,a Evgeniya V. Nurieva,*a
Elena R. Milaeva,a,b Sergei A. Kuznetsovc and Olga N. Zefirovaa,b
a Department of Chemistry, M. V. Lomonosov Moscow State University, 119991 Moscow, Russian Federation.
E-mail: E.Selunina@org.chem.msu.ru
b Institute of Physiologically Active Compounds, Russian Academy of Sciences, 142432 Chernogolovka,
Moscow Region, Russian Federation. E-mail: olgaz@med.chem.msu.ru
c Institute of Biological Sciences, University of Rostock, D-18059 Rostock, Germany.
E-mail: sergei.kuznetsov@uni-rostock.de
DOI: 10.1016/j.mencom.2021.05.003
Bicyclic isothioureas of N-(4-methoxyphenyl)-2-aminocyclo-
alkane[d]thiazole type were obtained using intramolecular
OMe
OMe
ꢀ
electrophilic
cyclization
of
N-(cycloalk-2-enyl)-N'-
S
S
n
N
N
(4-methoxyphenyl)thioureas. Isothiourea fused with seven-
membered ring caused noticeable changes of the morphology
of human lung carcinoma cells A549, but without affecting
their microtubule net.
N
N
ꢀ
R
R
n ꢀ
n
Cycloheꢁtaꢂfused derivative causes
total change of morꢁhology of lung
carcinoma cells Aꢃꢄꢅ
Keywords: thiazolin-2-amines, cycloalka[d]thiazoles, thioureas, isothioureas, heterocyclization, verubulin, lung carcinoma A549,
cancer cell morphology.
In loving memory of our Great Teacher academician Nikolay S. Zefirov
Interchange in aromatic residues is a classical approach used in
drug design for optimization of characteristics of the initial
molecule.1–4 The replacement of aromatic moiety by non-
aromatic one is applied more rarely due to the pronounced
differences in their geometric and electronic parameters and a
consequent loss of activity. In some cases, however, such
replacement allows one to enhance the activity (owing to
acquired ability to fill more completely the available space in the
target binding site) or to improve the pharmacochemical
parameters of the initial molecule.5 Literature search reveals
examples of successive replacement of aromatic group (thiazole,
pyridine, etc.) by cyclic isothiourea moiety in the design of
inhibitors of menin, mixed lineage leukemia fusion protein
interaction,6 ligands of g-aminobutyric acid receptors7 and
inhibitors of enzyme nitric oxide synthase.8 In the present work,
we carried out an analogous replacement in the structure of
verubulin.9 The latter is a potent (but not clinically approved)10
ligand of colchicine binding site of cell protein tubulin, a
validated molecular target of anticancer drugs.11–15 The proposed
modification was aimed to study the potential of cyclic
isothoureas as tubulin-targeted antimitotic agents.
in our previous works.16–19 First, a convenient procedure
involving sequential reactions of allylic bromination,
nucleophilic substitution and addition to appropriate anisidine
was fulfilled without evaporation and isolation of intermediate
highly volatile bromides and isothiocyanates 2a–c. In this way,
substituted thioureas 3a–c, 3'a–c were obtained as racemic
mixtures with acceptable yields. 1H NMR spectra of these
compounds contain the signal for CHN-cycloalkenyl proton at
OMe
NCS
i, ii
iii
H
40–75%
(16% for 3'a)
N
N
n
n
R
1a–c
2a–c
S
n
3a–c, 3'a–c
a n = 3
b n = 2
c n = 1
3a–c R = Me
3'a–c R = H
Scheme 1 Reagents and conditions: i, NBS, AIBN, CCl4, reflux, 1.5 h;
ii, KSCN, MeOH, 0–22 °C, 2 h; iii, 4-MeOC6H4N(R)H, DIPEA, CH2Cl2,
22 °C, 12 h.
Schemes 1–3 demonstrate the synthesis of target compounds
utilizing an intramolecular cyclization of cycloalkenyl-
substituted ureas as the key step under the conditions elaborated
OMe
OMe
H
S
i
3a,b; 3'b
N
N
4a n = 3, R = Me, 57%
4b n = 2, R = Me, 90%
4'b n = 2, R = H, 96%
R
nH
HBr
4a,b; 4'b
N
N
N
Me
Me
Verubulin
Scheme 2 Reagents and conditions: i, HBr, CH2Cl2, 22 °C, 24 h.
© 2021 Mendeleev Communications. Published by ELSEVIER B.V.
on behalf of the N. D. Zelinsky Institute of Organic Chemistry of the
Russian Academy of Sciences.
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Mendeleev Commun., 2021, 31, 288–290
ꢀaꢁ
ꢀbꢁ
ꢀcꢁ
OMe
ii
OMe
Br
H
H
H
S
S
i
3c, 3'c
N
N
N
N
R
R
HBr
5, 5'
R = Me, 50%
H
6, 6'
10 ꢂm
R = Me, 71%
6
5
5' R = H, 71%
6' R = H, 69%
Figure 1 Immunofluorescence microscopy of the cells A549 treated for
24 h with (a): 0.5% DMSO (normal MTs); (b) 10 µm of 4a (normal MTs,
cell contraction); (c) 100 µm of 4a (normal MTs, rounded cells).
Scheme 3 Reagents and conditions: i, Br2, CH2Cl2, 22 °C, 24 h; ii, Bu3SnH,
AIBN, toluene, 100 °C, 10 h.
4.9–5.5 ppm and the peaks for cycloalkenyl olefinic protons at
5.3–5.7 and 6.7–5.9 ppm (for the synthetic details and all
characteristics of novel compounds, see Online Supplementary
Materials).
other than tubulin. It is worth mentioning that the positive
features of the lead molecule Verubulin include its ability to
cause tumour vascular disruption and to inhibit the growth of
multidrug resistant cells.10 Hence, theoretically its structural
analogue 4a may maintain these properties, which makes
interesting further studies of novel cyclic isothiourea in the
experiments in vitro and in vivo.
Thioureas 3a,b and 3'b were converted into corresponding
fused bicyclic isothoureas 4a,b and 4'b using electrophilic
addition and simultaneous intramolecular cyclization under mild
conditions by treatment with AcBr/MeOH (see Scheme 2). The
cyclization occurred diastereoselectively with the formation of
products with cis-configuration in accordance with our earlier
study.16 In 1H NMR spectra of 4a,b and 4'b, the two resonances
in the range of 3.8–3.9 and 4.1–4.5 ppm correspond to the
chemical shifts of protons at the ring junction of the fused system,
while the signals at 60–67 and 48–52 ppm in 13C NMR spectra
refer to the appropriate carbon atoms. The peak of thiazole C2
atom is observed in the range of 169–174 ppm. Spectral data of
compounds 4a,b and 4'b indicate the presence of one tautomer,
however they are not diagnostic for exact elucidation of its
structure. It should be mentioned, that we failed to isolate an NH
analogue of 4a with seven-membered ring, because the reaction
with thiourea 3'a led to a complex mixture of products.
This work was supported by Russian Science Foundation
(project no. 19-13-00084). Part of the work, comprising the data
for compounds 3'b, 4'b, 5', was performed within the State
assignment AAAA-A16-116032250004-2. The authors
acknowledge B. Wobith (Rostock University) for technical
assistance, Yu. K. Grishin (Moscow University) for performance
of NMR experiments and the German organization DAAD for
support of academic co-operation between Moscow (State) and
Rostock Universities.
Online Supplementary Materials
Supplementary data associated with this article can be found
in the online version at doi: 10.1016/j.mencom.2021.05.003.
To synthesize the target isothioureas fused with five-
membered rings, the cyclization of thioureas 3c, 3'c was carried
out in the presence of bromine as more active electrophile (see
Scheme 3). The chromatographic purification gave individual
diastereoisomers 5, 5' (one signal sets were observed in their
1H NMR spectra). In 1H NMR spectrum of compounds 5, 5', the
peak for C6H–Br proton was observed at ~4.3 ppm and in
13C NMR spectrum, the corresponding carbon atom resonated in
the range of 60–63 ppm. The configuration of bromine atom was
assigned as trans towards thiazol ring on the basis of 1H NOESY
experiment (see Online Supplementary Materials). Bromo
substituted derivatives 5, 5' were subjected to reductive
debromination by treatment with tri-n-butyltin hydride to give
the target compounds 6, 6' (mass spectral data: [M+H]+ = 263
for 6 and 249 for 6'). The disappearance of the peaks at
~4.1–4.3 ppm in 1H NMR and at ~70 ppm in 13C NMR confirms
the removal of C6-bromine atom from initial compounds 5, 5'.
Analogously to 4, compounds 5 and 6 were obtained as single
tautomers.
Isothioureas 4a,b, 4'b and 6, 6' were studied for their ability
to change the morphology and microtubule net of human lung
carcinoma cells A549 using the standard procedures.20–22 The
compounds with cyclohexane and cyclopentane rings 4b, 4'b
and 6, 6' independently in the presence or absence of N-methyl
substituent did not show any effect even at a high concentration
of 100 mm. Only cyclohepta-fused isothiourea 4a demonstrated
noticeable changes in cell morphology (cell contraction and
rounding) giving evidence to its cytotoxic action both at 10 and
100 µm (Figure 1). Though compound 4a caused the termination
of the cell growth, no changes in microtubules (MTs) were
observed.
References
1 S. R. Langdon, P. Ertl and N. Brown, Mol. Inf., 2010, 29, 366.
2 G. A. Patani and E. J. LaVoie, Chem. Rev., 1996, 96, 3147.
3 N. A. Meanwell, J. Med. Chem., 2011, 54, 2529.
4 The Practice of Medicinal Chemistry, 2nd edn., ed. C. G. Wermuth,
Academic Press, Amsterdam, Boston, 2003.
5 F. Lovering, J. Bikker and Ch. Humblet, J. Med. Chem., 2009, 52, 6752.
6 J. Grembecka, S. He, A. Shi, T. Purohit, A. G. Muntean, R. J. Sorenson,
H. D. Showalter, M. J. Murai, A. M. Belcher, T. Hartley, J. L. Hess and
T. Cierpicki, Nat. Chem. Biol., 2012, 8, 277.
7 A. Giraudo, J. Krall, F. Bavo, B. Nielsen, K. T. Kongstad, B. Rolando,
R. De Blasio, D. E. Gloriam, R. Löffler, L. Thiesen, K. Harpsøe,
K. Frydenvang, D. Boschi, P. Wellendorph, M. L. Lolli, A. A. Jensen
and B. Frølund, J. Med. Chem., 2019, 62, 5797.
8 S. Ueda, H. Terauchi, A. Yano, M. Matsumoto, T. Kubo, Y. Kyoya,
K. Suzuki, M. Ido and M. Kawasaki, Bioorg. Med. Chem., 2004, 12, 4101.
9 N. Sirisoma, A. Pervin, H. Zhang, S. Jiang, J. A. Willardsen,
M. B. Anderson, G. Mather, C. M. Pleiman, S. Kasibhatla, B. Tseng,
J. Drewe and S. X. Cai, J. Med. Chem., 2009, 52, 2341.
10 Sh.-B. Wang, X.-F. Wang. B. Qin, E. Ohkoshi, K.-Y. Hsieh. E. Hamel,
M.-T. Cui, D.-Q. Zhu, M. Goto, S. L. Morris-Natschke, K.-H. Lee and
L. Xie, Bioorg. Med. Chem., 2015, 23, 5740.
11 E. C. McLoughlin and N. M. O’Boyle, Pharmaceuticals, 2020, 13, 8.
12 I. A. Gracheva, E. S. Shchegravina, H.-G. Schmalz, I. P. Beletskaya and
A. Yu. Fedorov, J. Med. Chem., 2020, 63, 10618.
13 L.-Y. Xia, Y.-L. Zhang, R. Yang, Zh.-Ch. Wang, Y.-D. Lu, B.-Zh. Wang
and H.-L. Zhu, Curr. Med. Chem., 2019, 26, 1.
14 K. Haider, S. Rahaman, M. Sh. Yar and A. Kamal, Expert Opin. Ther.
Pat., 2019, 29, 623.
15 G. La Regina, A. Coluccia, V. Naccarato and R. Silvestri, Eur. J. Pharm.
Sci., 2019, 131, 58.
16 E. V. Nurieva, T. P. Trofimova, A. A. Alexeev, A. N. Proshin,
E. A. Chesnakova, Yu. K. Grishin, K. A. Lyssenko, M. V. Filimonova,
S. O. Bachurin and O. N. Zefirova, Mendeleev Commun., 2018, 28, 390.
17 A. A. Alexeev, E. V. Nurieva, T. P. Trofimova, E. A. Chesnakova,
Yu. K. Grishin, K. A. Lyssenko, M. V. Filimonova and O. N. Zefirova,
Mendeleev Commun., 2019, 29, 14.
Therefore, one may suggest that the antimitotic action of
isothiourea 4a relates to the interaction with molecular target
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Mendeleev Commun., 2021, 31, 288–290
18 A. A. Alexeev, E. V. Nurieva, K. A. Lyssenko, Yu. K. Grishin and
O. N. Zefirova, Struct. Chem., 2019, 30, 473.
19 E. V. Nurieva, A. A. Alexeev, Yu. K. Grishin, V. A. Tafeenko and
21 N. A. Zefirov, Yu. A. Evteeva, A. I. Krasnoperova, A. V. Mamaeva,
E. R. Milaeva, S. A. Kuznetsov and O. N. Zefirova, Mendeleev
Commun., 2020, 30, 421.
O. N. Zefirova, Mendeleev Commun., 2020, 30, 145.
22 E. V. Nurieva, N. A. Zefirov, N. Fritsch, E. R. Milaeva, S. A. Kuznetsov
20 A. Al-Haddad, M. A. Shonn, B. Redlich, A. Blocker, J. K. Burkhardt,
H. Yu, J. A. Hammer, III, D. G. Weiss, W. Steffen, G. Griffiths and
S. A. Kuznetsov, Mol. Biol. Cell, 2001, 12, 2742.
and O. N. Zefirova, Mendeleev Commun., 2020, 30, 706.
Received: 20th February 2021; Com. 21/6458
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