Bioorganic & Medicinal Chemistry Letters 27 (2017) 792796

Contents lists available at ScienceDirect

Bioorganic & Medicinal Chemistry Letters

journal homepage: www.elsevier.com/locate/bmcl

A novel piperazine linked b-amino alcohols bearing a benzosuberone

scaffolds as anti-proliferative agents
Sowmya Vanguru a, Lavanya Jilla a, Yasodakrishna Sajja a, Rajashaker Bantu a, Lingaiah Nagarapu a,,
Jagadeesh Babu Nanubolu b, Bala Bhaskar c, Nishant Jain c, Sreekanth Sivan d, Vijjulatha Manga d
a
Organic Chemistry Division II (CPC), CSIR-Indian Institute of Chemical Technology, Tarnaka, Hyderabad 500007, India
b
Center for X-ray Crystallography, CSIR-Indian Institute of Chemical Technology, Tarnaka, Hyderabad 500007, India
c
Center for Chemical Biology, CSIR-Indian Institute of Chemical Technology, Tarnaka, Hyderabad 500007, India
d
Molecular Modeling and Medicinal Chemistry Group, Department of Chemistry, Osmania University, Tarnaka, Hyderabad 500007, India

a r t i c l e i n f o a b s t r a c t

Article history: A new series of 1-((9-chloro-2,3-dimethyl-6,7-dihydro-5H-benzo[7]annulen-8-yl)methoxy)-3-(4-phe-

Received 9 November 2016 nylpiperzin-1-yl) propan-2-ols (6a-k) have been designed, synthesized and their structures were estab-
Revised 30 December 2016 lished by spectroscopic data (FT-IR, 1H NMR, 13C NMR, HRMS) and further confirmed by X-ray analysis.
Accepted 11 January 2017
The newly synthesized compounds 6a-k were evaluated for their in vitro anti-proliferative activity
Available online 13 January 2017
against four cancer cell lines such as HeLa (cervical), MDA-MB-231 (breast), A549 (lung) and MIAPACA
(pancreatic). Among the compounds tested, the compound 6e displayed most potent activity against four
Keywords:
cancer cell lines with GI50 values ranging from 0.010 to 0.097 lM. The structure and anti-proliferative
Benzosuberone
Piperazine
activity relationship was further supported by in silico molecular docking study of the active compounds
b-Amino alcohol against Colchicine binding site of b-tubulin.
Anti-proliferative 2017 Elsevier Ltd. All rights reserved.
Cell lines
Molecular modeling

Introduction alcohol fragment is a common structural subunit in natural prod-

Cancer is one of the most common diseases worldwide; it is the
second-leading cause of deaths in the world. The identification of
new therapies is an area ongoing importance in biomedical
research.1,2 Cervical, breast, lung, colon, and pancreatic cancers
are most common in the developing and under developed coun-
tries. Therefore, the development of new anticancer agents and
more selective treatment strategies for cancer has received more
and more attention for medicinal chemists.
Natural products containing seven membered ring fused to an
aromatic ring attracted considerable attention in recent years
due to their remarkable biological activities. Benzocycloheptenone
and its derivatives are an important class of heterocyclic com-
pounds, which constitute the key core of various natural products
and play a unique role in drug discovery program. The benzo-
suberone moiety is the main scaffold of several natural products.
For example, Theaflavin3 present in black tea and Colchicine4 from
the plant Colchicum autumnale are two natural alkaloids with
potent anti-cancer activity (Fig. 1). On the other hand b-amino
Corresponding author.
E-mail addresses: lnagarapuiict@yahoo.com, nagarapu@iict.res.in (L. Nagarapu).
ucts and plays a key role in medicinal chemistry, pharmaceuticals
and in organic synthesis.5,6 Many b-amino alcohols exhibit a broad
spectrum of biological activities.7 These molecules are usually pre-
pared from aminolysis of epoxide and are non volatile, completely
odorless products. b-amino alcohols Hapalosin, Swainsonine are
found as potential anticancer agents (Fig. 1).811 Piperazine based
b-amino alcohols are known for their biological activity. They find
application as positive ionotropic agents, increasing myocardial
contractivity, in the treatment of cardiac disorders such as conges-
tive heart failure.1215 Some of the b-amino alcohols bearing the
piperazine motif (MS-073 and MS-209) (Fig. 1) are found to have
application in the reversal of multidrug resistance in cancer cells.16
In view of these observations and in continuation to our ongo-
ing research activities,17 to discover and develop tumor growth
inhibitors and apoptotic inducers as potential new anti-cancer
agents, we have made an attempt to design and synthesize three
biologically versatile heterocyclic scaffolds like benzosuberone,
b-amino alcohol linkage and piperazine in single molecular
platform (Fig. 2). The synthesized compounds were evaluated for
their in vitro anti-proliferative activity against four cancer cell lines
such as HeLa (cervical), MDA-MB-231 (breast), A549 (lung) and
MIAPACA (pancreatic) and a SRB cell proliferation assay was used
to estimate viability or growth. Furthermore, molecular docking

http://dx.doi.org/10.1016/j.bmcl.2017.01.031
0960-894X/ 2017 Elsevier Ltd. All rights reserved.
 S. Vanguru et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 792796
Fig. 1. Representative natural products bearing benzosuberone ring and b-amino
alcohols bearing the piperazine motif (MS-073 and MS-209).
Fig. 2. Design strategy for novel piperazine based b-amino alcohols of benzo-
suberone derivatives.
studies helped in revealing the mode of action of these compounds
through their interactions with the colchicine binding site of
b-tubulin.
To begin with, 2,3-dimethyl-6,7,8,9-tetrahydro benzocyclohep-
tenone (1)17 under go Vilsmeir-Haack-Arnold reaction by treating
with POCl3, dimethylformamide at 060 C for 4 h to afford the
(Z)-9-chloro-2,3-dimethyl-6,7-dihydro-5H-benzo[7]annulene-8-
carbaldehyde (2). The chloro aldehyde 2 was reduced by using
sodium borohydride in MeOH to afford corresponding
(9-chloro-2,3-dimethyl-6,7-dihydro-5H-benzo[7]annulene-8yl)
methanol (3). The key intermediate 2-(((9-chloro-2,3-dimethyl-
6,7-dihydro-5H-benzo[7]annulen-8-yl) methoxy) methyl)oxirane
(4) was obtained in 80% yield from the reaction of alcohol 3 with
()-epichlorohydrin in presence of THF/NaH at reflux temperature
for 3 h. The targeted novel 1-((9-chloro-2,3-dimethyl-6,7-dihydro-
5H-benzo[7]annulen-8-yl)methoxy)-3-(4-phenylpiperzin-1-yl)
propan-2-ols (6a-k) were achieved by treatment of 2-(((9-chloro-
2,3-dimethyl-6,7-dihydro-5H-benzo[7]annulen-8-yl) methoxy)
methyl)oxirane (4) with a variety of piperazines 5a-k in methanol
at reflux temperature in excellent yields (Scheme 1) and their
793
Scheme 1. Synthesis of b-amino alcohols of benzosuberone derivatives (6a-k).
1
structures were established by spectroscopic data (FT-IR, H
NMR, 13C NMR, HRMS).
The formation of compound 6a was evident from the ESI-MS
spectrum with the appearance of [M+H]+ peak at m/z 455 [HRMS
455.24530], while the FT-IR spectrum revealed the presence of a
broad absorption band at 3428 cm 1 which attributed to the pres-
ence of alcohol group, and the presence of ether linkage group in
6a was resonated at 1232 cm 1. Further, in IR spectrum, absorption
band at 2855 cm 1 indicated that secondary alcohol attached sp3
C-H group in 6a. Similarly, the 1H NMR spectrum of 6a exhibited
multiplet at d 3.964.05 due to secondary alcohol attached CH
proton. To provide further evidence for the proposed structure,
13
C NMR was recorded, which showed peak at d 60.7 due to sec-
ondary alcohol attached carbon, in between secondary alcohol
794 S. Vanguru et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 792796

and 6j showed promising anti-proliferative activity with GI50 val-

Fig. 3. ORTEP diagram of 6a (BA36) with the atom-numbering scheme. Displace-
ment ellipsoids are drawn at the 30% probability level and H atoms are shown as
small spheres of arbitrary radius (CCDC 1511980).
and piperazine CH2 carbon showed peak at d 66.0, whereas ether
linkage containing carbons showed peaks at d 71.4, 72.3 respec-
tively. Among these compounds, crystal structure of compound
6a was determined by X-ray diffraction analysis.18,19 Fig. 3 gives
a perspective view of 6a (BA36): which was in agreement with
the compound.
Effects of the compounds on the viability of human cancer cells
ues ranging from 0.1 to 2.9, 1.5 to 2.6, 0.010 to 0.097 and 1.2 to
2.9 lM respectively, against the four human cancer cell lines.
Among them, compound 6e showed potent anti-proliferative activ-
ity against all the four human cancer cell lines. In particularly com-
pound 6e showed potent anti-proliferative activity against three
cancer cell lines such as MDA-MB-231 (breast), A549 (lung) and
MIAPACA (pancreatic) with GI50 values 0.010, 0.023, and
0.056 lM respectively, which has better activity than standard
drug Nocodazole. In HeLa cell line compound 6e showed activity
with GI50 value 0.097 lM which was closer to the standard drug
Nocodazole. On the other hand in HeLa and MIAPACA cancer cell
lines compounds 6b and 6j showed moderate activity with GI50
values 1.0, 1.4, 1.5, and 1.2 lM. In case of A549 cancer cell line
compounds 6b and 6c showed moderate activity with GI50 values
0.9 and 1.5 lM.
According to this data, it has been clearly indicated that the
compound with ANO2 group on phenyl ring attached to the piper-
azine moiety i.e. 6e showed better anticancer activity than the
other compounds. It clearly indicated that the electron withdraw-
ing group such as nitro group played a key role towards difference
in biological activity.

All the synthesized compounds of novel b-amino alcohols 6a-k
were screened for their in vitro anti-proliferative activity against
four cancer cell lines such as HeLa (cervical), MDA-MB-231
(breast), A549 (lung) and MIAPACA (pancreatic) summarized in
Table 1.
The compounds were picked for an advanced assay against
these four human cancer cell lines at five concentrations (0.01,
0.1, 1, 10, 100 lM). GI50 (growth inhibitory activity) was calculated
and these values corresponded to the concentration of the com-
pound causing 50% decrease in the net cell growth as compared
with the standard drug, Nocodazole. Results were calculated for
each of these parameters if the level of activity was reached; how-
ever, if the effect was not achieved, the value was expressed as
greater or less than the maximum or minimum concentration
tested.
Based on Table 1, the synthesized compounds 6a-k showed
moderate to significant cancer cell growth inhibition with GI50
values ranging from 0.010 to 24.4 lM. The effect of various sub-
stituents on the benzosuberone tethered piperazine moiety was
examined. The structure-activity relationship (SAR) study revealed
that the orientation of substituent on piperazine ring is not only
crucial but also required benzosuberone along with a b-amino
alcohol linker for inducing the anti-proliferative activity against
these cancer cell lines. In particular, the compounds 6b, 6c, 6e
Molecular docking studies
Docking is a computational technique that places a small mole-
cule (ligand) in the binding site of its macromolecular target and
estimates its binding affinity.2026 In order to understand the
molecular interaction between the synthesized molecules and b-
tubulin, molecular docking followed by MM-GBSA calculations
was performed. Crystal structure of b-tubulin co crystallized with
nocodazole was downloaded from protein data bank (PDB id:
5CA1). GLIDE 5.6 was used for docking all the molecules into
5CA1, the change in binding free energy of protein ligand complex
was calculated by PRIME MM-GBSA protocol, where in the amino
acid residues with 5 radius of binding site were allowed to flex.
Each molecule protein complex was analyzed for interactions with
the receptor. Nocodazole showed three hydrogen bond interac-
tions (Fig. 4) with a binding affinity DGbind of 66.782 kcal/mol.
The synthesized molecules showed majorly hydrophobic interac-
tion with the active site amino acid residues, molecule 6e the best
active molecule in the series showed better binding affinity (DGbind
of 108.626 kcal/mol) than Nocodazole corroborating with exper-
imental anti-proliferative studies (GI50), the DGbind values are
given in Table 2. Nocodazole binds to Colchicine binding site of
b-tubulin through three hydrogen bonds with Glu 198 and Asn
165, the imidazole ring shows p-p interaction with phenyl ring

Table 1
GI50a values of the synthesized b-amino alcohols 6a-k against four human cancer cell lines.

Compounds HeLa MDA-MB-231 A549 MIAPACA

6a 6.9 0.8 7.8 0.7 5.8 0.9 8.8 0.7
6b 1.0 0.1 2.9 0.7 0.9 0.9 1.5 0.3
6c 1.7 0.7 2.1 0.8 1.5 0.6 2.6 0.8
6d 3.8 1.8 2.5 0.7 3.0 0.3 2.0 0.5
6e 0.097 0.7 0.010 0.7 0.023 0.9 0.056 0.8
6f 7.2 0.7 7.3 0.9 4.0 0.1 7.9 0.6
6g 16.3 0.2 20.8 0.5 18.2 0.3 24.4 0.6
6h 7.4 0.6 7.9 0.6 5.2 0.7 5.6 0.3
6i 7.8 0.9 9.0 0.5 6.3 0.6 9.6 0.7
6j 1.4 0.1 2.9 + 0.8 1.8 0.2 1.2 0.5
6k 2.4 0.1 3.0 0.7 2.1 0.6 2.8 0.9
Nocodazoleb 0.086 0.007 0.065 0.005 0.09 003 0.091 0.008

Bold values shown significant activity.

a
GI50: 50% Growth inhibition, concentration of drug (in lM) resulting in 50% reduction in net protein increase compared with control cells.
b
Positive controls.
 S. Vanguru et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 792796 795

Fig. 4. Binding interaction of Nocodazole in the Colchicine binding site of b-tubulin.

Fig. 6. Binding interaction of compound 6e in the Colchicine binding site of
b-tubulin.

Table 2
Binding affinity of molecules obtained from Prime MM-
In conclusion, we have designed and synthesized a novel piper-
GBSA calculation.
azine based b-amino alcohols of benzosuberone derivatives 6a-k in
Molecule DGbind (kcal/mol) excellent yields and their structures were confirmed by spectral
6a 88.181 analysis. Newly synthesized derivatives were evaluated for their
6b 97.004 in vitro anti-proliferative activity against four cancer cell lines such
6c 91.807 as HeLa, MDA-MB-231, A549 and MIAPACA. Among them com-
6d 52.116
6e 108.626
pounds 6b, 6c, 6e and 6j were showed better anti-proliferative
6f 79.621 activity with GI50 values ranging from 0.010 to 24.4 lM compared
6g 100.625 to other evaluated compounds. Particularly, the compound 6e has
6h 93.796 shown potent activity (GI50: 0.0100.097 lM) compared to stan-
6i 82.042
dard drug Nocodazole. From docking studies it could be assumed
6j 87.957
6k 85.160 that the synthesized molecules bind to the colchine binding site.
Nocodazole 66.782 Compound 6e has shown better binding affinity compared to
nocodazole.
Bold value shown significant activity.

of Tyr 200, the benzimidazole ring and thiophene ring occupies the
hydrophobic cavity of the binding site. The synthesized molecules
also bound to the colchicines binding site in a similar manner.
Fig. 5 shows the overlay of molecule 6e and Nocodazole in the
binding site. Molecular interaction of molecule 6e (Fig. 6)
benzocyloheptene ring occupying the hydrophobic cavity showed
alkyl-alkyl hydrophobic interaction with amino acid residues Leu
246, Lys 252, Ala 314 and Lys 350. Electrostatic interactions were
seen with Glu 198 and Try 250. The nitro phenyl group showed
p-p interaction with phenyl ring of Phe 167. Based on the above
observations it can be concluded that 6e has a better binding
affinity compared to Nocodazole.
Fig. 5. Dockpose overlay of Nocodazole (green) and compound 6e (blue) in the
Colchicine binding site of b-tubulin.
Acknowledgements
The authors gratefully acknowledge the financial support
through the project: DST-SERB/EMEQ-078/2013.
Supplementary material
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2017.01.
031.
References
1. Hatnapure GD, Keche AP, Rodge AH, Birajdar SS, Tale RH, Kamble VM. Bioorg
Med Chem Lett. 2012;22:6385.
2. Khan Z, Bisen PS. Biophys Res Acta. 2013;123:183.
3. Yang CS, Lambert JD, Ju J, Lu G, Sang S. Toxicol Appl Pharmacol. 2007;2:265.
4. Chaplin DJ, Hills SA. Int Radiat Oncol Biol Phys. 2002;54:1491.
5. Gallego L, Arroyo P. J Sep Sci. 2003;26:947.
6. Klingler FD. Acc Chem Res. 2007;40:1367.
7. (a) Angelina MA, Thiago N, Bianca SF, et al. Bioorg Med Chem Lett.
2013;23:2883;
(b) Fabien A, Bernard T, Eric H, Daniel C, Alain C, Sandrine B. J Chem Soc, Dalton
Trans. 2010;39:8982;
(c) Imzil L, Elamrani K, Charaf A, Bouzoubaa M, Leclerc G. J Pharm Belg.
1993;48:335.
8. (a) Hermann C, Pais GCG, Geyer A, Kuhnert SM, Maier ME. Tetrahedron.
2000;56:8461;
(b) Palomo C, Oiarbide M, Garcia JM, et al. J Org Chem. 2004;69:4126.
9. Lee HK, Chun JS, Pak CS. J Org Chem. 2003;68:2471.
10. Au CWG, Pyne SG. J Org Chem. 2006;71:7097.
11. Ceccon J, Greene AE, Poisson J-F. Org Lett. 2006;8:4739.
12. Press JB, Falotico R, Hajos ZG, et al. J Med Chem. 1992;35:4509.
13. Barton DL, Press JB, Hajos ZG, Sawyers RA. Tetrahedron Asymmetry. 1992;31189.
14. Butrous GS, Debbas NMG, Erwin J, et al. Eur Heart J. 1988;9489.
15. Sircar I, Haleen SJ, Burke S. Eur J Med Chem. 1992;35:4442.
16. Suzuki T, Fukazawa N, San-nohe K. J Med Chem. 1997;40:2047.
17. (a) Nagarapu L, Gaikwad HK, Sirikonda K, et al. Eur J Med Chem. 2010;45:4720;
(b) Nagarapu L, Gaikwad HK, Bantu R, Manikonda SR. Eur J Med Chem.
2011;46:2152;
(c) Nagarapu L, Vanaparthi S, Bantu R, Kumar CG. Eur J Med Chem. 2013;69:817;
796 S. Vanguru et al. / Bioorganic & Medicinal Chemistry Letters 27 (2017) 792796
(d) Nagarapu L, Yadagiri B, Bantu R, Kumar CG, Pombala S, Nanubolu J. Eur J Med
Chem. 2014;71:91;
(e) Yadagiri B, Holagunda BUD, Bantu R, et al. Eur J Med Chem. 2014;79:260;
(f) Bandi Y, Gurrala S, Bantu R, et al. Bioorg Med Chem Lett. 2015;25:2220;
(g) Bandi Y, Holagunda UD, Bantu R, et al. Bioorg Med Chem Lett. 2014;24:5041;
(h) Bollu R, Palem JD, Bantu R, Nagarapu L. Eur J Med Chem. 2015;89:138;
(i) Banu S, Bollu R, Bantu R, Nagarapu L. Eur J Med Chem. 2017;125:400.
18. SMART & SAINT. Software Reference Manuals. Versions 6.28a & 5.625. Madison,
Wisconsin, U.S.A.: Bruker Analytical X-ray Systems Inc.; 2001.
19. Sheldrick GM. SHELXS97 and SHELXL Version 2014/7. <http://shelx.uni-ac.gwdg.
de/SHELX/index.php>.
20. Kavallaris M. Nat Rev Cancer. 2010;10:194.
21. Zhou J, Giannakakou P. Curr Med Chem Anticancer Agents. 2005;5:65.
22. Hammond JW, Cai D, Verhey KJ. Curr Opin Cell Biol. 2008;20:71.
23. Schrdinger LLC Glide, Version 5.6. Prime Version 3.0, New York, NY.
24. Wang Y. FEBS J. 2016;283:102.
25. Friesner RA, Banks JL, Murphy RB, et al. J Med Chem. 2004;47:1739.
26. Hayes JM, Archontis G. J R Soc Interface. 2011;8:171190.