Synthesis and Antileukemic Activity of Novel 2-(4-(2,4-dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)ethanone Derivatives

Article abstract of DOI:10.1111/j.1747-0285.2011.01307.x

A series of novel 2-(4-(2,4-dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl) piperidin-1-yl)ethanone derivatives 9(a-e) and 10(a-g) were synthesized and characterized by ¹H NMR, IR, mass spectral, and elemental analysis. These novel co

Full text of DOI:10.1111/j.1747-0285.2011.01307.x

ª 2011 John Wiley & Sons A/S
doi: 10.1111/j.1747-0285.2011.01307.x
Chem Biol Drug Des 2012; 79: 360–367
Research Letter
Synthesis and Antileukemic Activity of Novel
2-(4-(2,4-dimethoxybenzoyl)phenoxy)-1-(4-(3-
(piperidin-4-yl)propyl)piperidin-1-yl)ethanone
Derivatives
Kambappa Vinaya1,2, Chandagirikoppal V.
tackle this disease (1,2). The search for pharmacological approaches
to neoplastic disease has made some impressive gains started after
1940 when the antileukemic activity of nitrogen mustard was dis-
covered during World War II. Among the modifications to the family
of antitumor compounds, heterocyclic compounds have been exten-
sively applied by many groups to modify the reactivity profile.
Kavitha3,4, Doddakunche S. Prasanna1,5
Siddappa Chandrappa1, Somasagara R.
,
Ranganatha1, Sathees C. Raghavan3 and
Kanchugarakoppal S. Rangappa1,*
1Department of Studies in Chemistry, University of Mysore,
Manasagangotri, Mysore 570 006, India
Benzophenone moiety-containing derivatives showed interesting cyto-
toxicity profiles (3,4). A wealth of data shows that the great impor-
tance of benzophenones is fundamentally because of the diverse
biological (5) and chemical (6) properties that they possess. Benzo-
phenone derivatives possess antimycobacterial (7–10), inhibitors of
HIV-1 reverse transcriptase (RT) (11,12) and anticancer (13) activities.
Recently, the in vitro effects of the benzophenone derivatives
garcinol, isogarcinol, and xanthochymol on cell growth in four human
leukemia cell lines were reported (14). Combretastatin A-4, a benzo-
phenone analogue, exhibits a selective toxicity for proliferating endo-
thelial cells in vitro by induction of apoptosis and vascular shut down
in human cancer models in vivo (4). Piperidine derivatives in general
are known to possess a diverse range of biological activity. A piperi-
dine ring is part of an antitumor drug such as CPT-11 (15). Com-
pounds containing piperidine moiety play an important role in the
field of medicinal chemistry. Derivatives of this class possess useful
biological activities such as herbicidal, insecticidal, fungicidal, bacte-
ricidal, anti-inflammatory, antihistaminic, hypotensive, anticancer,
CNS stimulant and depressant, and nerve activities (16–23).
2Department of Chemistry, Government First Grade College, Kadur
577 548, India
3Department of Biochemistry, Indian Institute of Science, Bangalore
560 012, India
4Department of Chemistry, Government College for Women, Mandya
571 401, India
5Post-graduate Department of Studies in Chemistry, JSS College of
Arts, Commerce and Science, Ooty Road, Mysore 570 025, India
*Corresponding author: Kanchugarakoppal S. Rangappa,
rangappaks@chemistry.uni-mysore.ac.in; rangappaks@gmail.com
A series of novel 2-(4-(2,4-dimethoxybenzoyl)phen-
oxy)-1-(4-(3-(piperidin-4-yl)propyl) piperidin-1-yl)etha-
none derivatives 9(a–e) and 10(a–g) were synthesized
and characterized by 1H NMR, IR, mass spectral, and
elemental analysis. These novel compounds were
evaluated for their antileukemic activity against two
human leukemic cell lines (K562 and CEM) by using
the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazo-
liumbromide assay. Some of the tested compounds
showed good antiproliferative activity with IC50 val-
ues ranging from 1.6 to 8.0 lM. Compound 9c, 9e, and
10f with an electron-withdrawing halogen substitu-
ent at the para position on the phenyl ring showed
excellent in vitro potency against tested human leu-
kemia cells (K562 and CEM).
The literature has recorded several instances, in which incorporation
of two of the structural features required for activity in a single
molecule has given rise to significant enhancement in activity
(24,25). Because of the broad spectrum of activities reported in the
literature so far, we have synthesized a system that combines two
biolabile components together, targeting potent molecules possess-
ing antileukemic activity. Continuing the search for new compounds
with anticancer activity in the group of bioactive heterocycles, we
have undertaken the synthesis of title compounds and investigated
the antiproliferative activity against human leukemia cells.
Key words: 1,3-dimethoxy benzene, anticancer agents, cell death,
leukemia, MTT assay, piperidine
Received 23 December 2010, revised 18 November 2011 and accepted
for publication 1 December 2011
Experimental Section
In recent years, great progress has been made in the treatment of
cancer, and so the life expectation of patients with cancer has been
improved remarkably. Emerging science within the past decade has
created many opportunities for fundamentally new approaches to
Materials and methods
Infrared (IR) spectra were recorded using a Jasco FTIR-4100
spectrometer in the wave number range of 4000–400⁄ cm (Jasco, Dun-
360

Antileukemic Activity of Novel Derivatives
mow, UK). Nuclear magnetic resonance (1H NMR) spectra were
recorded on a Bruker AM 400 MHz spectrometer using DMSO-d6 as
solvent and tetramethylsilane as an internal standard. The chemical
shifts are expressed in d, and the following abbreviations are used: s
(singlet), d (doublet), t (triplet), q (quartet), and m (multiplet). Mass and
purity were recorded on a LC-MSD-Trap-XCT. Elemental (CHNS) analy-
ses were obtained on Vario EL III Elementar. The purity of the com-
pounds was checked by thin-layer chromatography (TLC). Silica gel
column chromatography was performed using Merck 7734 silica gel
(60–120 mesh) and Merck-made TLC plates. All the reagents and chem-
icals were from Sigma Aldrich Chemicals Pvt Ltd (Bangalore, India).
anone (6) (0.25 g, 0.491 mmol), 3-methoxyphenyl isocyanate (7b)
(0.073 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield:
82%. 1H NMR (DMSO-d6, 400 MHz) d: 9.22 (s, 1H, –CONH), 7.42
(d, 2H, Ar–H), 7.21 (d, 1H, Ar–H), 7.14 (t, 1H, Ar–H), 6.92 (m, 4H,
Ar–H), 6.81 (d, 1H, Ar–H), 6.63 (d, 2H, Ar–H), 5.25 (s, 2H, –OCH2),
3.85 (s, 3H, –OCH3), 3.78 (s, 3H, –OCH3), 3.73 (s, 3H, –OCH3), 2.83–
2.98 (m, 8H), 2.07 (bs, 2H), 1.09–1.25 (m, 14H). IR (KBr, cm)1):
3356, 2889, 1716, 1648, 1293, 1250, 1124, 1105. MS (ESI) m ⁄ z:
658.3 (M+H+). Anal. calcd. for C38H47N3O7 (in %): C-69.38, H-7.20,
N-6.39. Found C-69.56, H-7.38, N-6.26.
Synthesis of 4-(3-(1-(2-(4-(2,4-
General procedure for synthesis of 2-(4-(2,4-
dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-
yl)propyl)piperidin-1-yl)ethanone derivatives
9(a–e) and 10(a–g)
dimethoxybenzoyl)phenoxy)acetyl)piperidin-4-
yl) propyl)-N-(4-fluorophenyl)piperidine-1-
carboxamide 9c
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), 4-fluorophenyl isocyanate (7c)
(0.067 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield:
77%. 1H NMR (DMSO-d6, 400 MHz) d: 9.19 (s, 1H, –CONH), 7.79
(d, 2H, Ar–H), 7.27-7.33 (m, 2H, Ar–H), 6.94 (m, 4H, Ar–H), 6.86 (d,
1H, Ar–H), 6.54 (d, 2H, Ar–H), 5.25 (s, 2H, –OCH2), 3.87 (s, 3H, –
OCH3), 3.72 (s, 3H, –OCH3), 2.89–2.98 (m, 8H), 2.81 (bs, 2H), 1.74
(m, 2H), 1.09–1.26 (m, 14H). IR (KBr, cm)1): 3346, 2885, 1710, 1639,
1287, 1252, 1121, 1039. MS (ESI) m ⁄ z: 646.4 (M+H+). Anal. calcd.
for C37H44FN3O6 (in %): C-68.82, H-6.87, N-6.51. Found C-68.88, H-
6.82, N-6.63.
A solution of 2-(4-(2,4-dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-
4-yl)propyl) piperidin-1-yl)ethanone (6) (1.0 eq) in dry dichlorome-
thane was taken and cooled to 0–5 ꢀC in an ice bath. Triethylamine
(3.0 eq) was added to this cold reaction mixture and stirred for
10 min, and then different isocyanates 7(a–e) (1.0 eq) or isothiocy-
anates 8(a–g) (1.0 eq) were added and allowed to stir at room
temperature for 5–6 h. Progress of the reaction mixture was moni-
tored by TLC. Upon completion, the solvent was removed under
reduced pressure, and residue was taken in water and extracted
with ethyl acetate. The organic layer was washed with 10% ammo-
nium chloride solution, and finally, water wash was given to the
organic layer and dried with anhydrous sodium sulfate; the solvent
was evaporated to yield crude product that was purified by column
chromatography over silica gel (60–120 mesh) using hexane ⁄ ethyl
acetate (8:2) as eluent.
Synthesis of N-(3-chlorophenyl)-4-(3-(1-(2-(4-(2,4-
dimethoxybenzoyl)phenoxy)acetyl) piperidin-4-
yl)propyl)piperidine-1-carboxamide 9d
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), 3-chlorophenyl isocyanate (7d)
(0.075 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield:
Synthesis of 4-(3-(1-(2-(4-(2,4-
dimethoxybenzoyl)phenoxy)acetyl)piperidin-4-
yl)propyl) -N-(2-methoxyphenyl)piperidine-1-
carboxamide 9a
1
75%. H NMR (DMSO-d6, 400 MHz) d: 9.20 (s, 1H, –CONH), 7.84 (s,
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), 2-methoxyphenyl isocyanate (7a)
(0.073 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield:
80%. 1H NMR (DMSO-d6, 400 MHz) d: 9.17 (s, 1H, –CONH), 7.41
(d, 2H, Ar–H), 7.32 (m, 2H, Ar–H), 7.08 (m, 4H, Ar–H), 6.83 (d, 1H,
Ar–H), 6.62 (d, 2H, Ar–H), 5.21 (s, 2H, –OCH2), 3.84 (s, 3H, –OCH3),
3.76 (s, 3H, –OCH3), 3.73 (s, 3H, –OCH3), 2.81–3.0 (m, 8H), 2.05 (bs,
2H), 1.12–1.24 (m, 14H). IR (KBr, cm)1): 3356, 2889, 1716, 1648,
1293, 1250, 1124, 1105. MS (ESI) m ⁄ z: 658.3 (M+H+). Anal. calcd.
for C38H47N3O7 (in %): C-69.38, H-7.20, N-6.39. Found C-69.44, H-
7.26, N-6.36.
1H, Ar–H), 7.75 (d, 1H, Ar–H), 7.14 (t, 1H, Ar–H), 7.05 (d, 1H, Ar–
H), 6.90 (m, 4H, Ar–H), 6.83 (d, 1H, Ar–H), 6.59 (d, 2H, Ar–H), 5.22
(s, 2H, –OCH2), 3.83 (s, 3H, –OCH3), 3.72 (s, 3H, –OCH3), 2.80–2.93
(m, 8H), 2.07 (bs, 2H), 1.07–1.22 (m, 14H). IR (KBr, cm)1): 3361,
2920, 2862, 1728, 1674, 1278, 1256, 1123, 1045, 725. MS (ESI)
m ⁄ z: 662.3 (M+H+). Anal. calcd. for C37H44ClN3O6 (in %): C-67.11,
H-6.70, N-6.35. Found C-67.27, H-6.79, N-6.22.
Synthesis of N-(4-chlorophenyl)-4-(3-(1-(2-(4-(2,4-
dimethoxybenzoyl)phenoxy)acetyl) piperidin-4-
yl)propyl)piperidine-1-carboxamide 9e
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), 4-chlorophenyl isocyanate (7e)
(0.075 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield:
75%. 1H NMR (DMSO-d6, 400 MHz) d: 9.15 (s, 1H, –CONH), 7.81
(d, 2H, Ar–H), 7.25-7.30 (m, 2H, Ar–H), 6.97 (m, 4H, Ar–H), 6.85 (d,
1H, Ar–H), 6.59 (d, 2H, Ar–H), 5.27 (s, 2H, –OCH2), 3.85 (s, 3H, –
OCH3), 3.74 (s, 3H, –OCH3), 2.85–2.95 (m, 8H), 2.83 (bs, 2H), 1.78
Synthesis of 4-(3-(1-(2-(4-(2,4-
dimethoxybenzoyl)phenoxy)acetyl)piperidin-4-
yl) propyl)-N-(3-methoxyphenyl)piperidine-1-
carboxamide 9b
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
Chem Biol Drug Des 2012; 79: 360–367
361

Vinaya et al.
(m, 2H), 1.09–1.21 (m, 12H). IR (KBr, cm)1): 3361, 2920, 2862, 1728,
1674, 1278, 1256, 1123, 1045, 725. MS (ESI) m ⁄ z: 662.3 (M+H+).
Anal. calcd. for C37H44ClN3O6 (in %): C-67.11, H-6.70, N-6.35. Found
C-67.16, H-6.77, N-6.41.
Synthesis of N-(3-chlorophenyl)-4-(3-(1-(2-(4-
(2,4-dimethoxybenzoyl)phenoxy)acetyl)
piperidin-4-yl)propyl)piperidine-1-
carbothioamide 10d
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), 3-chlorophenyl isothiocyanate (8d)
(0.083 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield:
73%. 1H NMR (DMSO-d6, 400 MHz) d: 9.22 (s, 1H, –CONH), 7.74
(d, 1H, Ar–H), 7.40 (d, 2H, Ar–H), 7.12 (t, 1H, Ar–H), 6.91 (m, 4H,
Ar–H), 6.80 (d, 1H, Ar–H), 6.63 (d, 2H, Ar–H), 5.25 (s, 2H, –OCH2),
3.82 (s, 3H, –OCH3), 3.74 (s, 3H, –OCH3), 2.80–2.91 (m, 8H), 2.05
(bs, 2H), 1.08–1.20 (m, 14H). IR (KBr, cm)1): 2952, 2876, 1741,
1648, 1270, 1241, 1129, 1097, 722. MS (ESI) m ⁄ z: 678.3 (M+H+).
Anal. calcd. for C37H44ClN3O5S (in %): C-65.52, H-6.54, N-6.20.
Found C-65.58, H-6.59, N-6.26.
Synthesis of 4-(3-(1-(2-(4-(2,4-
dimethoxybenzoyl)phenoxy)acetyl)piperidin-4-
yl)propyl)-N-(2-methoxyphenyl)piperidine-1-
carbothioamide 10a
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), 2-methoxyphenyl isothiocyanate
(8a) (0.081 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol).
1
Yield: 75%. H NMR (DMSO-d6, 400 MHz) d: 9.14 (s, 1H, -CSNH),
7.33 (d, 2H, Ar–H), 7.25 (d, 1H, Ar–H), 7.20 (d, 1H, Ar–H), 7.05 (m,
4H, Ar–H), 6.93 (d, 1H, Ar–H), 6.62 (d, 2H, Ar–H), 5.25 (s, 2H, –
OCH2), 3.83 (s, 3H, –OCH3), 3.72 (s, 3H, –OCH3), 3.68 (s, 3H, –
OCH3), 2.85–2.98 (m, 8H), 2.05 (bs, 2H), 1.12–1.20 (m, 14H). IR (KBr,
cm)1): 2939, 2881, 1730, 1643, 1274, 1245, 1128, 1117, 1041. MS
(ESI) m ⁄ z: 674.2 (M+H+). Anal. calcd. for C38H47N3O6S (in %): C-
67.73, H-7.03, N-6.24. Found C-67.65, H-6.92, N-6.16.
Synthesis of 4-(3-(1-(2-(4-(2,4-
dimethoxybenzoyl)phenoxy)acetyl)piperidin-4-
yl)propyl) -N-(3,4,5-trimethoxyphenyl)piperidine-
1-carbothioamide 10e
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)
ethanone (6) (0.25 g, 0.491 mmol), 3,4,5-trimethoxyphenyl isothiocya-
nate (8e) (0.110 g, 0.491 mmol), and triethylamine (0.15 g,
Synthesis of 4-(3-(1-(2-(4-(2,4-
dimethoxybenzoyl)phenoxy)acetyl)piperidin-4-
yl)propyl) -N-(3-methoxyphenyl)piperidine-1-
carbothioamide 10b
1
1.47 mmol). Yield: 70%. H NMR (DMSO-d6, 400 MHz) d: 9.50 (s, 1H,
–CONH), 7.42 (d, 2H, Ar–H), 7.24 (d, 1H, Ar–H), 7.11 (t, 1H, Ar–H),
6.94 (m, 4H, Ar–H), 6.83 (d, 1H, Ar–H), 5.23 (s, 2H, –OCH2), 3.75 (s,
15H, –OCH3), 2.80–2.93 (m, 8H), 2.07 (bs, 2H), 1.09–1.21 (m, 14H). IR
(KBr, cm)1): 2959, 2880, 1735, 1653, 1276, 1240, 1124, 1058, 727.
MS (ESI) m⁄z: 734.4 (M+H+). Anal. calcd. for C40H51N3O8S (in %): C-
65.46, H-7.00, N-5.73. Found C-65.41, H-7.03, N-5.76.
The product obtained was pale yellow oily liquid from 2-(4-(2,4-
dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-
yl)ethanone (6) (0.25 g, 0.491 mmol), 3-methoxyphenyl isothiocya-
nate (8b) (0.081 g, 0.491 mmol), and triethylamine (0.15 g,
1.47 mmol). Yield: 75%. 1H NMR (DMSO-d6, 400 MHz) d: 9.25 (s,
1H, –CONH), 7.40 (d, 2H, Ar–H), 7.24 (d, 1H, Ar–H), 7.16 (t, 1H,
Ar–H), 6.94 (m, 4H, Ar–H), 6.83 (d, 1H, Ar–H), 6.66 (d, 2H, Ar–H),
5.27 (s, 2H, –OCH2), 3.83 (s, 3H, –OCH3), 3.75 (s, 3H, –OCH3),
3.70 (s, 3H, –OCH3), 2.80–2.93 (m, 8H), 2.07 (bs, 2H), 1.09–1.21
(m, 14H). IR (KBr, cm)1): 2939, 2881, 1730, 1643, 1274, 1245,
1128, 1117, 1041. MS (ESI) m ⁄ z: 674.1 (M+H+). Anal. calcd. for
C38H47N3O6S (in %): C-67.73, H-7.03, N-6.24. Found C-67.80, H-
6.98, N-6.28.
Synthesis of N-(4-chlorophenyl)-4-(3-(1-(2-(4-
(2,4-dimethoxybenzoyl)phenoxy)acetyl)
piperidin-4-yl)propyl)piperidine-1-
carbothioamide 10f
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), 4-chlorophenyl isothiocyanate (8f)
(0.083 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield:
1
Synthesis of 4-(3-(1-(2-(4-(2,4-
74%. H NMR (DMSO-d6, 400 MHz) d: 9.21 (s, 1H, -CSNH), 7.82 (d,
dimethoxybenzoyl)phenoxy)acetyl)piperidin-4-
yl)propyl) -N-(2-fluorophenyl)piperidine-1-
carbothioamide 10c
2H, Ar–H), 7.32 (m, 2H, Ar–H), 6.93 (m, 5H, Ar–H), 6.57 (d, 2H, Ar–
H), 5.25 (s, 2H, –OCH2), 3.85 (s, 3H, –OCH3), 3.73 (s, 3H, –OCH3),
2.85–2.92 (m, 8H), 2.08 (bs, 2H), 1.10–1.19 (m, 14H). IR (KBr, cm)1):
2952, 2876, 1741, 1648, 1270, 1241, 1129, 1097, 722. MS (ESI)
m ⁄ z: 678.3 (M+H+). Anal. calcd. for C37H44ClN3O5S (in %): C-65.52,
H-6.54, N-6.20. Found C-65.58, H-6.59, N-6.26.
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), 2-fluorophenyl isothiocyanate (8c)
(0.075 g, 0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield:
1
77%. H NMR (DMSO-d6, 400 MHz) d: 9.23 (s, 1H, -CSNH), 7.82 (d,
2H, Ar–H), 7.37 (d, 2H, Ar–H), 7.11 (m, 4H, Ar–H), 6.95 (d, 1H, Ar–
H), 6.65 (d, 2H, Ar–H), 4.75 (s, 2H, –OCH2), 3.87 (s, 3H, –OCH3),
3.72 (s, 3H, –OCH3), 1.75 (m, 4H), 1.09–1.35 (m, 16H), 0.91 (m, 4H).
IR (KBr, cm)1): 2892, 1717, 1632, 1280, 1252, 1125, 1041. MS (ESI)
m ⁄ z: 662.2 (M+H+). Anal. calcd. for C37H44FN3O5S (in %): C-67.15,
H-6.70, N-6.35. Found C-67.21, H-6.65, N-6.36.
Synthesis of 4-(3-(1-(2-(4-(2,4-
dimethoxybenzoyl)phenoxy)acetyl)piperidin-4-yl)
propyl)-N-phenylpiperidine-1-carbothioamide 10g
The product obtained was pale yellow oily liquid from 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) (0.25 g, 0.491 mmol), phenyl isothiocyanate (8g) (0.066 g,
362
Chem Biol Drug Des 2012; 79: 360–367

Antileukemic Activity of Novel Derivatives
1
0.491 mmol), and triethylamine (0.15 g, 1.47 mmol). Yield: 70%. H
ble cells to metabolize a yellow tetrazolium salt to violet formazan.
Exponentially growing K562 or CEM cells (1 · 104 cells ⁄ well) were
plated in triplicates and incubated with 5, 10, and 20 lM of 9(a–e)
and 10(a–g). Cells were harvested after 48 and 72 h of treatment
and incubated with MTT (5 mg ⁄ mL) at 37 ꢀC. The blue MTT forma-
zan precipitate was then solubilized in detergent (50% final concen-
tration of N,N-dimethylformamide and 10% of sodium dodecyl
sulfate). Absorbance was measured at 570 nm using ELISA plate
reader (Molecular Devices, USA, Sunnyvale, CA, USA). The mean
absorbance of culture medium was used as the blank and was sub-
tracted. IC50 values (concentration of compound causing 50% inhibi-
tion of cell growth) were estimated after 72 h of compound
treatment. The absorbance of vehicle cells was taken as 100% viabil-
ity, and the values of treated cells were calculated as a percentage
of control and presented as histograms (Figures 1 and 2).
NMR (DMSO-d6, 400 MHz) d: 9.21 (s, 1H, –CONH), 7.16 (m, 5H,
Ar–H), 6.90 (m, 4H, Ar–H), 6.83 (d, 1H, Ar–H), 6.64 (d, 2H, Ar–H),
5.25 (s, 2H, –OCH2), 3.81 (s, 3H, –OCH3), 3.73 (s, 3H, –OCH3), 2.80–
2.91 (m, 8H), 2.05 (bs, 2H), 1.09–1.21 (m, 14H). IR (KBr, cm)1):
2892, 1717, 1632, 1280, 1252, 1125, 1041. MS (ESI) m ⁄ z: 644.7
(M+H+). Anal. calcd. for C37H45N3O5S (in %): C-69.02, H-7.04, N-
6.53. Found C-68.95, H-7.00, N-6.48.
In vitro anti-proliferative activity
Cell lines and culture
Human cell line, K562 (chronic myelogenous leukemia) and CEM
were purchased from National Center for Cell Science, Pune, India.
Cells were grown in RPMI 1640 containing 10% FBS, 100 U of
Penicillin G ⁄ mL, and 100 lg of streptomycin ⁄ mL at 37 ꢀC in a
humidified atmosphere containing 5% CO2.
Results and Discussion
Chemistry
MTT assay
For the synthesis of the target key intermediate compound 6,
the reaction sequences outlined in Scheme 1 were followed. (4-
Hydroxyphenyl)(2,4-dimethoxyphenyl) methanone (3) was synthes-
Cell survival was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl
tetrazolium bromide (MTT) assay, which is based on the ability of via-
Figure 1: Determination of the effect of 2-(4-(2,4-dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)ethanone derivatives
9(a–e) and 10(a–g) on cell proliferation by MTT assay. After 48 and 72 h of exposure, K562 cells with 9(a–e) and 10(a–g) (5, 10 and
20 lM) were incubated with MTT (5 mg ⁄ mL) in duplicates, and resulting blue formazan precipitate was dissolved in detergent, and absor-
bance was measured at 570 nm. Results are presented as percentage of cell proliferation (the cell viability of vehicle cells were considered
as 100%).
Chem Biol Drug Des 2012; 79: 360–367
363

Vinaya et al.
ised by Friedel-Crafts reaction with 1,3-dimethoxy benzene (1)
(1.0 eq) and p-hydroxy benzoic acid (2) (1.2 eq) in the presence
of phosphorus oxychloride (7.0 eq) and zinc chloride (2.5 eq) at
60–70 ꢀC for 2 h. The absence of –COOH proton peak in 1H
NMR spectra confirmed the formation of compound 3. Treatment
of (4-hydroxyphenyl)(2,4-dimethoxyphenyl) methanone (3) with
chloroacetic acid (4.5 eq) in potassium carbonate (7.0 eq) solu-
tion and refluxed for 10 h gave the O-alkylated product. The
absence of Ar–OH and presence of –COOH proton peak in com-
pound 4 confirm the formation of the product. 2-(4-(2,4-Dim-
ethoxybenzoyl) phenoxy)acetic acid (4) (1.0 eq) and 4-(3-(piperidin-
4-yl)propyl)piperidine (5) (1.0 eq) in N,N-dimethyl formamide
(DMF) were taken, to which N-methylmorpholine (NMP) (3.0 eq)
and 10% of o-benzotriazol-1-yl-N,N,N1,N1-tetramethyl uranium tet-
rafluoroborate (TBTU) catalyst were added, and reaction mixture
was stirred for 5 h at room temperature, which gave target key
intermediate 6. The absence of –COOH proton peak and pres-
ence of –NH proton peak confirmed the formation of compound
6. The nucleophilic substitution reaction of 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)eth-
anone (6) with different substituted aromatic isocyanates 7(a–e)
(R–N=C=O) ⁄ isothiocyanates 8(a–g) (R-N=C=S) was carried out in
the presence of triethylamine and dichloromethane as solvent
with a good yield of 74–80%. The absence of –NH and pres-
ence of –CO–NH, –CS–NH proton peak in synthesized derivatives
9(a–e), and 10(a–g) in 1H NMR spectra confirmed the identity
of the products. It is also confirmed by IR data, for carboxamide
series 9(a–e) and thioamide series 10(a–g); IR data showed
stretching frequency at 3350–3360 ⁄ cm for –NH and 1640–
1660 ⁄ cm for –C=O group. The chemical structures of all the syn-
thesized compounds are given in Table 1.
Biology
Compounds 9(a–e) and 10(a–g) were evaluated for their cytotoxic
activity in vitro against human leukemia cells, K562 (chronic myeo-
logenous leukemia) and CEM. We used MTT assay to assess the
effect of 2-(4-(2,4-dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-
yl)propyl)piperidin-1-yl)ethanone derivatives 9(a–e) and 10(a–g) on
cell viability. For this, cells growing in the log phase were treated
with different concentrations (5, 10, and 20 lM) of 2-(4-(2,4-dim-
ethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)
piperidin-1-
Figure 2: Determination of the effect of 2-(4-(2,4-dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl)piperidin-1-yl)ethanone derivatives
9(a–e) and 10(a–g) on cell proliferation by MTT assay. After 48 and 72 h of exposure, CEM cells with 9(a–e) and 10(a–g) (5, 10 and
20 lM) were incubated with MTT (5 mg ⁄ mL) in duplicates, and resulting blue formazan precipitate was dissolved in detergent, and absor-
bance was measured at 570 nm. Results are presented as percentage of cell proliferation (the cell viability of vehicle cells were considered
as 100%).
364
Chem Biol Drug Des 2012; 79: 360–367

Antileukemic Activity of Novel Derivatives
Scheme 1: 2-(4-(2,4-Dimethoxybenzoyl)phenoxy)-1-(4-(3-(piperidin-4-yl)propyl) piperidin-1-yl)ethanone derivatives.
yl)ethanone derivatives 9(a–e) and 10(a–g). To ensure that the
solvent per se had no effect on the cell growth, negative control
tests were performed using DMSO at the concentration used for
highest concentration of the compound tested.
with a methoxy group (9a and 9b) resulted in five- and threefold
reduced activity with IC50 values 8.0 and 5.5 lM, respectively.
Among thiourea derivatives 10(a–g), compounds 10d and 10f
showed good inhibition with IC50 value of 2.3 and 2.1 lM, respec-
tively. The remaining compounds in the series showed two- to
threefold less inhibitory activity. The inhibition by compound 10d
and 10f could be attributed to the electron-withdrawing chloro
groups present on the substituted thiourea at the meta and para
positions, respectively.
The effects of the compounds were expressed as percentage of cell
proliferation. The IC50 value was determined for 72 h, and the
results are summarized in Table 1. As shown in Table 1, most of
the compounds exhibited inhibition of the growth of both K562 and
CEM cells in the micromolar range. From the Figures 1 and 2, it
was found that incorporation of 4-fluorophenyl (9c) and 4-chlor-
ophenyl (9e) resulted interesting inhibition with IC50 values of 2.2
and 1.6 lM, respectively, for K562 cells and 2.3 and 2.0 lM,
respectively, for CEM cells. Among the urea analogs 9(a–e),
compounds 9c, 9d, and 9e showed good inhibition with IC50 val-
ues 2.2, 2.4, and 1.6 lM, respectively, for K562 cells. Within the
few variations considered in this study, the presence of electron-
withdrawing groups afforded a clear beneficial effect with regard
to antiproliferative properties. However, the replacement of chloro
Conclusion and Future Directions
In conclusion, this study demonstrates that the synthesized com-
pounds exhibit interesting anti-leukemia properties. The synthesized
compounds exhibited good antiproliferative activity with IC50 values
ranging from 1.6 to 8.0 lM. Further studies are required to know
the mechanism of action of these molecules. These studies are cur-
rently under progress.
Chem Biol Drug Des 2012; 79: 360–367
365

Vinaya et al.
Table 1: Chemical structure and IC50 values of the synthesized
compounds 9(a–e) and 10(a–g) as determined based on MTT
assay
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Acknowledgments
We are grateful to UGC, Govt. of India, for financial support to KSR
and KV under the projects vide no. F. 39-106 ⁄ 2010 (SR) and F. 39-
810 ⁄ 2010 (SR). SCR acknowledges support from Lady Tata Memo-
rial Trust international award for leukemia research (London).
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