Synthesis and biological evaluation of 2-amino-7,7-dimethyl 4-substituted-5-oxo-1-(3,4,5-trimethoxy)-1,4,5,6,7,8-hexahydro-quinoline-3-carbonitrile derivatives as potential cytotoxic agents

Article abstract of DOI:10.1016/j.bmcl.2009.10.065

A large number of antimitotic drugs, derived from natural sources or chemically synthesized, have been identified and shown to interfere with the tubulin system. Inhibition of tubulin polymerization is among the important targets useful in the cancer ther

Full text of DOI:10.1016/j.bmcl.2009.10.065

Bioorganic & Medicinal Chemistry Letters 19 (2009) 6939–6942
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of 2-amino-7,7-dimethyl
4-substituted-5-oxo-1-(3,4,5-trimethoxy)-1,4,5,6,7,8-hexahydro-quinoline-
3-carbonitrile derivatives as potential cytotoxic agents
Saleh I. Alqasoumi a, Areej M. Al-Taweel b, Ahmed M. Alafeefy c, Mostafa M. Hamed d, Eman Noaman e,
Mostafa M. Ghorab a,
*
a Medicinal, Aromatic and Poisonous Plants Research Center (MAPPRC), College of Pharmacy, King Saud University, PO Box 2457, Riyadh 11451, Saudi Arabia
b Pharmacognosy Department, of Pharmacy, King Saud University, PO Box 2457, Riyadh 11451, Saudi Arabia
c Medicinal Chemistry Department, College of Pharmacy, King Saud University, PO Box 2457, Riyadh 11451, Saudi Arabia
d Department of Pharmaceutical chemistry, Faculty of Pharmacy & Biotechnology, German University in Cairo, New Cairo City, Egypt
e Radiation Biology Department, National Center for Radiation Research and Technology, Atomic Energy Authority, Nasr City, Cairo, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
A large number of antimitotic drugs, derived from natural sources or chemically synthesized, have been
identified and shown to interfere with the tubulin system. Inhibition of tubulin polymerization is among
the important targets useful in the cancer therapy.
The present work reports the synthesis of some novel quinoline derivatives bearing a trimethoxyphenyl
moiety. The trimethoxybenzene moiety has been reported to be crucial to obtain relevant cytotoxic and
antitubulin responses. All the newly synthesized compounds were evaluated for their in vitro anticancer
activity. Several compounds showed interesting cytotoxic activities compared to the used reference drug.
Ó 2009 Elsevier Ltd. All rights reserved.
Received 7 August 2009
Revised 11 October 2009
Accepted 15 October 2009
Available online 20 October 2009
Keywords:
Hexahydroquinoline
Trimethoxyphenyl
Anticancer
Tubulin
In the development of compounds that can be used in antican-
cer chemotherapy, the mitotic spindle was found to be among the
attractive targets. The mitotic spindle is formed by microtubules
which are cykloskeletal polymers of tubulin involved in many cel-
lular functions, and the compounds affecting the spindle will lead
to the accumulation of cells arrested in metaphase.1–4 Several anti-
mitotic drugs with diverse structures, derived from natural sources
or obtained by screening compound libraries, have been identified
and shown to affect the tubulin system.4–6
One of the most important tubulin-binding agents is combre-
tastatin A-4 (I), isolated from the bark of the South African tree
Combretum caffrum,7 strongly inhibits the polymerization of tubu-
lin by binding to the colchicine site.8 This compound had partic-
ular interest not only for its anticancer activity but also because it
has been from the simplest natural compounds with potent anti-
tubulin activity,8 and so SAR studies have been done to evaluate a
large number of developed combretastatin A-4 analogues.9
Among synthetic small-molecule tubulin inhibitors, was the CA-
4 analogue named phenstatin (II). Recently a series of thiophene
derivatives (III) have been synthesized and were found to be
potent inhibitors of tubulin polymerization.10 Romagnoli et al.
have then synthesized a series of benzo[b]thiophene (IV) and tet-
rahydro[2,3-c]pyridine (V) derivatives showing interesting activi-
ties in inhibiting microtubule polymerization and cell
proliferation (Fig. 1).4,11
In addition, quinoline derivatives have received considerable
attention because of their pivotal role in various biological pro-
cesses and numerous derivatives of quinolines have been reported
to have wide biological activities including the anticancer activ-
ity.12–16 Therefore, in a search for novel cytotoxic agents with an
expected antimitotic activity, we prompted to synthesize a new
series of hexahydroquinoline derivatives bearing a trimethoxy-
phenyl moiety. This trimethoxyphenyl moiety has been reported
to be crucial to obtain relevant cytotoxic and antitubulin potency
for the series of molecules occupying the colchicines-binding site
in the tubulin protein.4,9,17 All the newly synthesized compounds
were evaluated for their in vitro anticancer activity against the
used reference drug.
The present work reports the possible utility of 3-(3,4,5-tri-
methoxyanilino)-5,5-dimethyl-cyclohex-2-enone 3 in the synthe-
sis of 2-amino-7,7-dimethyl-5-oxo-4-substituted-aryl-1-(3,4,5-
trimethoxyphenyl)-1,4,5,6,7,8-hexahydro-quinoline-3-carbonitr-
iles 6a–t.
* Corresponding author. Tel.: +966 534292860; fax: +966 (01) 4670560.
E-mail address: mmsghorab@yahoo.com (M.M. Ghorab).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.10.065

6940
S. I. Alqasoumi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6939–6942
Figure 1. Different compounds which inhibit tubulin polymerization.
Enaminone 3 was obtained by condensation of 5,5-dimethyl-
cyclohexan-1,3-dione 1 with 3,4,5-trimethoxyaniline 2 (Scheme
1). The structure of compound 3 was supported by elemental anal-
ysis and spectral data. IR spectrum of compound 3 revealed the
presence of bands for NH at 3270 cmÀ1, for (CH aliph.) at 2955,
2930, 2824 cmÀ1 and for C@O at 1620 cmÀ1. Also, 1H NMR spec-
trum (in DMSO-d6), indicated the presence of a signal at 8.7 ppm
which could be assigned to NH of enaminone 3 and 3.6, 3.71,
3.79 ppm for the three methoxy groups.
We have studied the reaction of activated cyano olefins 4a–t
with enaminone 3 as a route to functionally substituted quinolines.
Condensation of 3 with 4a–t in a molar ratio (1:1) in refluxing eth-
anol containing triethylamine afforded the hexahydroquinoline
derivatives 6a–t. Formation of compounds, 6a–t, could be ex-
plained via initial Michael addition of enaminone 3 to the ylidenic
bond in 4a–t forming an acyclic intermediate 5a–t which cyclizes
by nucleophilic attack of the NH group on the cyano carbon, fol-
lowed by tautomerisation to the final products 6a–t.
Compounds 6a–t were unambiguously synthesized by another
route involving one-pot condensation of the appropriate alde-
hydes, malononitrile and enaminone 3 in a molar ratio (1:1:1) in
refluxing ethanol containing triethyamine (TEA) as catalyst. In this
case, formation of 6a–t are illustrated in terms of initial condensa-
tion of the aldehyde with malononitrile affording the activated
cyano olefin 4a–t, followed by addition of the enaminone 3 to
arylidenemalononitrile 4a–t as above.
The N-aryl substituent, 3,4,5-trimethoxyphenyl, promoted the
nucleophilicity of the enaminone 3 towards 2-arylidenemalononit-
rile 4a–t. Compound 3 nucleophilically adds to the arylidenemal-
ononitrile 4a–t to form an intermediate iminium ion that is
deprotonated by TEA (Scheme 2). The structure of compounds
6a–t was established on the basis of analytical and spectral data.
Thus, IR spectra of compounds 6a–t showed bands at 3469–
3206 cmÀ1 due (NH2), 2226–2162 cmÀ1 (CN), 1668–1620 cmÀ1
(C@O).
Scheme 1.
Doxorubicin, the reference drug used in this study is one of the
most effective antitumor agents used to produce regressions in
acute leukemia’s Hodgkin’s disease, and other lymphomas. The
relationship between survival ratio and drug concentration was
plotted to obtain the survival curve of Ehrlich Ascites Carcinoma
(EAC) cell line. The response parameter calculated was IC50 value
(Table 1), which corresponds to the compound concentration caus-
ing 50% mortality in net cells.
All the newly synthesized compounds were evaluated for their
in vitro cytotoxic activity. Compounds 6f containing 2-methoxy-
phenyl at position-4 with cyano group at position-3 (IC50 va-
lue = 26.58
cyano group at position-3 (IC50 value = 52.85
4-nitrophenyl at position-4 with cyano group at position-3 (IC50
lM), 6b having 4-methylphenyl at position-4 with
lM), and 6k bearing

S. I. Alqasoumi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6939–6942
6941
Figure 2. Docking pose of compound 6f ‘red’ in the colchicine site. DAMA-
colchicine in green.
In order to validate our docking procedure, we docked the
DAMA-colchicine which is the ligand already co-crystallised with
the protein. The docking results showed that the best scored con-
formation exhibits a similar fashion as the crystallized conforma-
tion with an RMSD of 1.1 Å.
Scheme 2. Postulated mechanism for the formation of compounds 6a–t.
We then docked the most active compounds, 6f and 6k, and in
both cases, it was found that the trimethoxyphenyl moiety is posi-
tioned in a different place when compared to the corresponding
colchicinoid A ring, and even the whole molecule exhibits totally
different conformation than the colchicine analogue in the crystal-
lized protein complex. (Figs. 2 and 3)
value = 49.60
ence drug doxorubicin ((IC50 value = 68.13
pound 6d having 4-hydroxyphenyl at position-4 with cyano group
at position-3 (IC50 value = 75.78 M) is nearly as active as doxoru-
l
M) are more potent and efficacious than the refer-
l
M). Additionally com-
l
bicin. On the other hand compounds 6m, 6p and 6s are less active
From the figures, it is clear that the compounds showing signif-
icant cytotoxic activity exhibits a different conformation and bind-
ing mode, in the colchicine binding site of tubulin, compared to the
DAMA-colchicine. Therefore, it could be suggested that these com-
pounds may exert their cytotoxic activity by a different mechanism
of action other than being antimitotic agents through the inhibi-
tion of the tubulin polymerization.
than doxorubicin as positive control.
A molecular modeling study has been performed to investigate
the possible binding conformation for this group of compounds to
the colchicine binding site of tubulin, which may give a suggestion
about their proposed mechanism of action as antitubulin agents.
Compounds 6fand 6k were docked in the (PDB entry: 1SA0)3 using
MOE 2007.09 (MOE).18
In conclusion, the objective of the present study was to synthe-
size and investigate the anticancer activity of new hybrid com-
pounds comprising hexahydroquinolines, bearing cyano group at
position-3 with different moieties at position-4 and 3,4,5-trime-
thoxyphenyl at position-1, for synergistic purpose. The results re-
vealed that the corresponding quinoline derivatives 6f, 6b and 6k
showed the highest in vitro cytotoxic activity when compared to
other tested compounds and doxorubicin as a reference drug.
Table 1
In vitro cytotoxic activity of the newly synthesized compounds 6a–t
a
a
Compound
Non-viable cells (%)
Concentration (
g mLÀ1
IC50
(lg/mL)
IC50 (lM)
l
)
100
50
25
10
Doxorubicin
100
10
90
10
100
40
100
15
20
40
30
80
40
100
45
40
90
30
40
50
20
68
8
60
2
30
6
50
0
20
30
65
1
15
30
20
50
20
55
10
30
10
5
24
0
30
0
10
10
30
0
10
10
5
10
10
20
2
20
2
37
68.13
—
52.85
—
75.78
—
26.58
—
6a
6b
6c
6d
6e
6f
6g
6h
6i
>100*
25
>100*
36
90
30
70
11
20
40
30
70
30
95
40
30
15
10
40
41
15
>100*
13
>100*
>100*
>100*
>100*
25
—
—
—
6j
6k
6l
49.60
—
94.69
—
>100*
50
6m
6n
6o
6p
6q
6r
6s
6t
>100*
>100*
75
—
142.85
0
>100*
>100*
100
—
—
30
31
10
10
21
5
215.98
—
>100*
a
IC50 value: corresponds to the compound concentration causing 50% mortality
in net cells.
Figure 3. Docking pose of compound 6k ‘red’ in the colchicine site. DAMA-
colchicine in green.
*
Compounds with IC50 >100 lg/mL are considered to be inactive.

6942
S. I. Alqasoumi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6939–6942
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