Synthesis and cytotoxic activity of certain trisubstituted azetidin-2-one derivatives as a cis-restricted combretastatin A-4 analogues

Article abstract of DOI:10.1007/s12272-016-0849-y

Novel series of 1,3,4-trisubstituted azetidin-2-one derivatives 8a–p were synthesized and proposed as cytotoxic agents acting via inhibition of tubulin at the colchicine binding site. The design of the target compounds was based upon modification in the s

Full text of DOI:10.1007/s12272-016-0849-y

Arch. Pharm. Res.
DOI 10.1007/s12272-016-0849-y
RESEARCH ARTICLE
Synthesis and cytotoxic activity of certain trisubstituted azetidin-
2-one derivatives as a cis-restricted combretastatin A-4 analogues
Salwa Elmeligie1 Azza. T. Taher1,2 Nadia A. Khalil1 Ahmed H. El-said3
•
•
•
Received: 11 April 2016 / Accepted: 12 October 2016
Ó The Pharmaceutical Society of Korea 2016
Abstract Novel series of 1,3,4-trisubstituted azetidin-2-
one derivatives 8a–p were synthesized and proposed as
cytotoxic agents acting via inhibition of tubulin at the
colchicine binding site. The design of the target compounds
was based upon modification in the structure of the vas-
cular targeting agent combretastatin A-4 (CA-4). The cis
double bond linker in CA-4 was replaced with the azetidin-
2-one ring aiming to prevent the cis/trans isomerization
that suppresses the activity of CA-4, thereby enhancing its
antiproliferative activity. All new compounds were inves-
tigated in vitro against MCF-7 and HCT-116 cell lines. The
inhibition of tubulin polymerization by four most potent
compounds 8g, 8j, 8n and 8o was also evaluated. The
synthesis of the final targets was achieved adopting Stau-
dinger reaction. Molecular modeling studies were per-
formed to rationalize the biological results.
Introduction
Antimitotic agents comprise one of the major classes in
chemotherapy, and microtubules are a significant target for
many natural anticancer agents (Jordan 2002). Micro-
tubules are the main constituents of the mitotic spindle and
are formed by association of protofilaments that are made
up of alternating a and b tubulin monomers. They are
involved in many essential functions throughout the cell
cycle including transport of material within the cell,
movement of the cell itself, and proper progression through
cell division (Downing 2000). Drugs acting as tubulin
inhibitors arrest the cell during mitosis, as they have the
ability to disrupt microtubule assembly or disassembly
making them preferential for tumor cells with high prolif-
eration rate (Dumontet and Jordan 2010).
Combretastatins are a group of tubulin-binding diaryl-
stilbenes isolated from the South African tree Combretum
caffrum. Combretastatin-A4 (CA-4) 2 comprises as one of
the most powerful antimitotic agents that inhibits the for-
mation of mitotic spindle through binding to tubulin at the
colchicine binding site, thereby interfering with cell growth
and proliferation (Dumontet and Jordan 2010; Cragg and
Newman 2005; Lippert 2007). In contrast to colchicine 1, the
anti-vascular effects of CA-4 in vivo are apparent well below
the maximum tolerated dose, offering a wide therapeutic
window (Tozer et al. 1999; West and Price 2004). A water
soluble pro-drug combretastatin A-4 phosphate (CA-4P) 3 is
under evaluation in phase 2 clinical trials for treatment of
non-small cell lung cancer and in phase 3 for treatment of
anaplastic thyroid cancer (O’Boyle et al. 2011a).
Keywords Tubulin Á Combretastatin A4 Á Azetidin-2-one Á
Cytotoxic activity
Electronic supplementary material The online version of this
article (doi:10.1007/s12272-016-0849-y) contains supplementary
material, which is available to authorized users.
& Salwa Elmeligie
salwaelmeligie@yahoo.com
1
Pharmaceutical Organic Chemistry Department, Faculty of
Pharmacy, Cairo University, Cairo, Egypt
2
Organic Chemistry Department, Faculty of Pharmacy,
October University for Modern Sciences and Arts (MSA),
6th October City, Giza, Egypt
Many conformationally restricted analogues of CA-4
have been reported (Wang et al. 2002; Ohsumi et al. 1998), in
which the isomerizable cis-double bond is replaced with a
heterocyclic ring, among which, the azetidin-2-one (b-
3
Pharmaceutical Organic Chemistry Department, Faculty of
Pharmacy, Misr University for Science and Technology,
6th October City, Giza, Egypt
123

S. Elmeligie et al.
lactam) ring comprises one of the important scaffolds
(O’Boyle et al. 2011b). The anticancer activity of some b-
lactam-containing compounds including potent non-iso-
merizable CA-4 analogues have been reported (Banik et al.
2003).
aldehyde 5 (100 mmol each) in ethanol (50 mL) containing
glacial acetic acid (2–3 drops) was heated under reflux for
5–12 h. After cooling, the crystalline solid product was
filtered off, then recrystallized from ethanol. Compounds
6a–e and 6g–i were prepared and purified as previously
reported (Al-Tai et al. 1976; Suresh et al. 2013; O’Boyle
et al. 2010; Klimczak et al. 2012; Greene et al. 2016).
Herein, it was of interest to synthesize certain new
combretastatin-related compounds containing the b-lactam
ring with the aim to provide rigidification of the structure to
prevent cis/trans isomerization observed in CA-4. The b-
lactam ring was also supposed to impart a useful scaffold
structure with a similar spatial arrangement between the
two phenyl rings as observed in the cis conformation of
CA-4. Moreover, in the newly synthesized compounds the
trimethoxyphenyl scaffold which is present in colchicine
and CA-4 was retained at N-1 or at C-4 of the azetidine-2-
one ring, that could afford additional sites of interaction
with the colchicine binding site.
N-(2-Bromobenzylidene)-3,4,5-trimethoxybenzenamine (6f)
Was prepared from 3,4,5-trimethoxybenzenamine and
2-bromobenzaldhyde applying the above procedure. Time
of reflux 10 h, Yield 83 %, pale yellow crystals, mp
135–136 °C. IR (KBr, t cm-1): 3061 (CH aromatic), 2999,
1
2938 (CH aliphatic), 1584 (C=N); H NMR (DMSO-d6)d
ppm: 3.67 (s, 3H, OCH3), 3.82 (s, 6H, 2OCH3), 6.63 (s, 2H,
Ar–H), 7.44–7.54 (m, 2H, Ar–H), 7.76 (d, 1H, Ar–H,
J = 7.5 Hz), 8.09 (d, 1H, Ar–H, J = 7.5 Hz), 8.78 (s, 1H,
CH=N). EIMS m/z (%rel. abundance): 349.00 (M-H,
99.98), 350.00 (M?, 29.59), 351.00 (M?H, 100), 352.00
(M?2, 27.63); Anal. Calcd for C16H16BrNO3: C 54.87, H
4.61, N 4.00. Found: C 55.12, H 4.70, N 4.18.
Materials and methods
Chemistry
General method for preparation of 8a–p
The starting materials 6a–e and 6g–i were prepared as
reported. Other chemicals and reagents were obtained from
Aldrich, Fluca or Merck and were used without further
purification unless otherwise indicated. Methylene chloride
was dried by distillation from calcium hydride prior to use.
Progress of the reactions was monitored using TLC sheets
precoated with UV fluorescent silica gel Merck 60F 254. The
solvent system was hexane and ethyl acetate (8:2) and spots
were visualized using UV lamp. IR spectra were determined
on Shimadzu FT-IR 8400 s spectrophotometer (KBr, cm-1).
1H NMR spectra were carried out using a Mercury 300-BB
300 MHz and Bruker 400 MHz spectrophotometers using
tetramethylsilane (TMS) as internal standard. 13C NMR
spectra were carried out using a Mercury 300-BB 75 MHz
and Bruker 100 MHz using TMS as internal standard.
Chemical shifts (d) are recorded in ppm on d scale, Micro-
analytical unit, Faculty of Pharmacy, Cairo University,
Egypt. Mass spectra were performed on Hewlett Packard
5988 spectrometer, Microanalytical Center, Cairo Univer-
sity, Egypt. Elemental analyses were carried out at the
Regional Center for Mycology and Biotechnology, Al-
Azhar University, Egypt. Melting points were determined on
Stuart apparatus and the values given are uncorrected. Col-
umn chromatography was carried out using standard silica
gel 60 (230–400 mesh) obtained from Fluca.
A mixture of the appropriate 6a–i (5 mmol) in anhydrous
CH2Cl2 (50 mL) and TEA (15 mmol, 1.08 mL) was heated
to reflux. A respective substituted acetyl chloride 7
(7.5 mmol) was injected drop wise through a rubber stop-
per over a period of 20 min, then heating was continued for
additional 7–12 h. After cooling, the reaction mixture was
washed with distilled water (50 mL 9 2) followed by
saturated aqueous NaHCO3 solution (50 mL), then with
10 % brine (50 mL). The organic layer was dried over
(anhyd. Na2SO4), and the solvent was distilled off under
reduced pressure to give the crude product, which was
further purified using column chromatography over silica
˚
gel 60 A (70–230 mesh) and (hexane/ethyl acetate gradi-
ent) as an eluent.
Microwave method for the preparation of 8l–o
To a solution of the appropriate 6g–i (5 mmol) in chol-
robenzene was added a respective acid chloride 7
(7.5 mmol) and TEA (15 mmol, 1.08 mL). The mixture
was irradiated in Milestone- StartSYNTH microwave oven
for 10 min at 80 °C, then washed with distilled water
(50 mL), saturated aqueous NaHCO3 solution (50 mL) and
brine (50 mL). The organic layer was dried (anhyd. Na2-
SO4), and concentrated under reduced pressure to give the
crude product, which was further purified using column
General method for preparation of Schiff bases (6a–i)
˚
chromatography over silica gel 60A (70–230 mesh) and
A solution containing equimolar amounts of the appropri-
ate substituted primary aromatic amine 4 and an aromatic
(hexane/ethyl acetate gradient) as an eluent.
123

Synthesis and cytotoxic activity of certain trisubstituted azetidin-2-one derivatives…
4-(2-Bromophenyl)-3-chloro-1-(4-methoxyphenyl)azetidin-
2-one (8a) Was prepared from N-(2-bromobenzylidene)-
4-methoxybenzenamine (6a) and acyl chloride 7. Yield
48 %, buff crystals, mp 109–110 °C. IR (KBr, t cm-1):
3062 (CH aromatic), 2945, 2829 (CH aliphatic), 1761
(C=O); 1H NMR (300 MHz, DMSO-d6)d ppm: 3.71 (s, 3H,
OCH3), 5.23 (d, 1H, H-4, J = 1.8 Hz), 5.57 (d, 1H, H-3,
J = 1.8 Hz), 6.94 (d, 2H, Ar–H, J = 9 Hz), 7.20 (d, 2H,
Ar–H, J = 9 Hz), 7.23 (d, 1H, Ar–H, J = 9.6 Hz),
7.31–7.38 (m, 2H, Ar–H), 7.75 (d, 1H, Ar–H, J = 9.6 Hz);
13C NMR (100 MHz, DMSO)d ppm: 55.76 (OCH3), 62.42
(C-3), 64.21 (C-4), 115.14 (2C), 119.05, 119.44 (2C),
128.35, 129.95 (2C), 131.26, 133.82, 134.32, 156.87 (ArC),
160.34 (C=O); EIMS m/z (%rel. abundance): 365.00 (M?,
84.52), 367.00 (M?2, 80.95); Anal. Calcd for C16H13-
BrClNO2: C 52.41, H 3.57, N 3.82. Found: C 52.51, H
3.60, N 3.89.
3-Chloro-4-(4-chlorophenyl)-1-(4-methoxyphenyl)-3-methyl
azetidin-2-one (8d) Was prepared from N-(4-chloroben-
zylidene)-4-methoxybenzenamine (6c) and acyl chloride 7.
Yield 74 %, pale brown crystals, mp 108–109 °C. IR (KBr,
t cm-1): 3028 (CH aromatic), 2987, 2839 (CH aliphatic),
1
1772 (C=O); H NMR (300 MHz, DMSO-d6)d ppm: 1.29
(s, 3H, CH3), 3.70 (s, 3H, OCH3), 5.63 (s, 1H, H-4), 6.91
(d, 2H, Ar–H, J = 9 Hz), 7.20 (d, 2H, Ar–H, J = 9 Hz),
7.36 (d, 2H, Ar–H, J = 8.4 Hz), 7.46 (d, 2H, Ar–H,
J = 8.4 Hz); 13C NMR (100 MHz, DMSO)d ppm: 20.76
(CH3), 55.72 (OCH3), 68.82 (C-3), 73.24 (C-4), 115.06
(2C), 119.66 (2C), 122.98 (2C), 129.34 (2C), 129.63,
132.36, 134.13, 156.82 (ArC), 162.71 (C=O); EIMS m/z
(%rel. abundance): 335.00 (M?, 58.47), 336.10 (M?H,
9.01). Anal. Calcd for C17H15Cl2NO2: C 60.73, H 4.50, N
4.17. Found: C 60.91, H 4.54, N 4.26.
4-(4-Chlorophenyl)-1-(4-methoxyphenyl)-3-(2-theinyl) aze-
tidin-2-one (8e) Was prepared from N-(4-chlorobenzyli-
dene)-4-methoxybenzenamine (6c) and acyl chloride 7.
Yield 54 %, buff crystals, mp 146–147 °C. IR (KBr, t
cm-1): 3010 (CH aromatic), 2958, 2837 (CH aliphatic),
4-(2-Bromophenyl)-3-chloro-1-(4-methoxyphenyl)-3-methy-
lazetidin-2-one (8b) Was prepared from N-(2-bromoben-
zylidene)-4-methoxybenzenamine (6a) and acyl chloride 7.
Yield 78 %, white crystals, mp 144–145 °C. IR (KBr, t
cm-1): 3070 (CH aromatic), 2970, 2841 (CH aliphatic),
1
1739 (C=O); H NMR (300 MHz, DMSO-d6)d ppm: 3.69
(s, 3H, OCH3), 4.70 (d, 1H, H-4, J = 2.7 Hz), 5.30 (d, 1H,
H-3, J = 2.7 Hz), 6.89 (d, 2H, Ar–H, J = 9 Hz),
6.91–7.15 (m, 4H, Ar–H), 7.23 (d, 2H, Ar–H, J = 9 Hz),
7.48–7.55 (m, 3H, Ar–H); 13C NMR (100 MHz, DMSO)d
ppm: 55.72 (C-3), 59.47 (OCH3), 62.67 (C-4), 114.99 (2C),
119.03 (2C), 126.49, 126.72, 127.84, 128.90 (2C), 129.52
(2C), 130.57, 133.67, 136.55, 136.62, 156.36 (ArC), 164.02
(C=O); EIMS m/z (%rel. abundance): 368.26 (M-H,
12.52), 369.22 (M?, 8.61); Anal. Calcd for C20H16ClNO2S:
C 64.95, H 4.36, N 3.79. Found: C 65.18, H 4.43, N 3.84.
1
1761 (C=O); H NMR (300 MHz, DMSO-d6)d ppm: 2.11
(s, 3H, CH3), 3.72 (s, 3H, OCH3), 5.74 (s, 1H, H-4), 6.96
(d, 2H, Ar–H, J = 9 Hz), 7.24 (d, 2H, Ar–H, J = 9 Hz),
7.30–7.36 (m, 3H, Ar–H), 7.76 (d, 1H, Ar–H, J = 9 Hz);
13C NMR (100 MHz, DMSO)d ppm: 24.88 (CH3), 55.79
(OCH3), 66.90 (C-3), 74.25 (C-4), 115.08 (2C), 115.13
(2C), 119.44, 119.71, 123.44, 128.20, 128.57, 130.07,
134.31, 156.84 (ArC), 163.28 (C=O); EIMS m/z (%rel.
abundance): 379.87 (M?, 2.87), 381.87 (M?2, 3.55),
383.87 (M?4, 0.87); Anal. Calcd for C17H15BrClNO2: C
53.64, H 3.97, N, 3.68. Found: C 53.78, H 3.99, N 3.68.
3-Chloro-4-(furan-2-yl)-1-(4-methoxyphenyl)azetidin-2-one
(8f) Was prepared from N-[(furan-2-yl)methylene]-4-
methoxybenzenamine (6e) and acyl chloride 7. Yield
64 %, brown oil. IR (KBr, t cm-1): 3001 (CH aromatic),
29254, 2927 (CH aliphatic), 1754 (C=O); 1H NMR
(300 MHz, DMSO-d6)d ppm: 3.70 (s, 3H, OCH3), 5.44 (s,
1H, H-4, J = 2.1 Hz), 5.47 (s, 1H, H-3, J = 2.1 Hz),
6.50–6.83 (m, 2H, furan H), 6.90 (d, 2H, Ar–H, J = 9 Hz),
7.26 (d, 2H, Ar–H, J = 9 Hz), 7.71 (d, 1H, furan H); 13C
NMR (100 MHz, DMSO)d ppm: 55.75 (OCH3), 58.17 (C-
3), 60.34 (C-4), 111.48, 112.41, 115.01 (2C), 119.26 (2C),
130.06, 145.02, 147.70, 156.90 (ArC), 160.32 (C=O);
EIMS m/z (%rel. abundance): 276.97 (M?, 18.35), 278.98
(M?2, 6.59); Anal. Calcd for C14H12ClNO3: C 60.55, H
4.36, N 5.04. Found: C 60.78, H 4.42, N 5.17.
4-(4-Bromophenyl)-3-chloro-1-(4-methoxyphenyl)azetidin-
2-one (8c) Was prepared from N-(4-bromobenzylidene)-
4-methoxybenzenamine (6b) and acyl chloride 7. Yield
45 %, white rod crystals, mp 148–149 °C. IR (KBr, t
cm-1): 3010 (CH aromatic), 2966, 2953 (CH aliphatic),
1
1751 (C=O). H NMR (300 MHz, DMSO-d6)d ppm: 3.69
(s, 3H, OCH3), 5.21 (d, 1H, H-4, J = 2.1 Hz), 5.37 (d, 1H,
H-3, J = 2.1), 6.88 (d, 2H, Ar–H, J = 9 Hz), 7.20 (d, 2H,
Ar–H, J = 9 Hz), 7.42 (d, 2H, Ar–H, J = 8.4 Hz), 7.60 (d,
2H, Ar–H, J = 8.4 Hz); 13C NMR (100 MHz, DMSO)d
ppm: 55.74 (OCH3), 62.61 (C-3), 64.19 (C-4), 115.06 (2C),
119.54, 122.90 (2C), 129.67 (2C), 129.76 (2C), 132.47,
134.91, 156.81 (ArC), 160.42 (C=O); EIMS m/z (%rel.
abundance): 364.84 (M?, 2.61), 366.85 (M?2, 3.51),
368.86 (M?4, 0.90); Anal. Calcd for C16H13BrClNO2: C
52.41, H 3.57, N 3.82. Found: C 52.49, H 3.61, N 3.90.
4-(3,4,5-Trimethoxyphenyl)-1-(4-methoxyphenyl)-3-(2-thienyl)-
azetidin-2-one (8g) Was prepared from N-(3,4,5-
trimethoxybenzylidene)-4-methoxybenzenamine (6d) and
123

S. Elmeligie et al.
acyl chloride 7. Yield 70 %, dark brown crystals, mp
130–131 °C. IR (KBr, t cm-1): 3003 (CH aromatic), 2927,
2839 (CH aliphatic), 1757 (C=O); H NMR (400 MHz,
7. Yield 39 %, buff crystals, mp 126–127 °C. IR (KBr, t
cm-1): 3001(CH aromatic), 2953, 2926 (CH aliphatic),
1
1
1743 (C=O); H NMR (300 MHz, DMSO-d6)d ppm: 3.60
DMSO-d6)d ppm: 3.67 (s, 3H, OCH3), 3.69 (s, 3H, OCH3),
3.77 (s, 6H, 2OCH3), 4.83 (d, 1H, H-4, J = 2 Hz), 5.18 (d,
1H, H-3, J = 2 Hz), 6.87 (s, 2H, Ar–H), 6.90–7.54 (m, 7H,
4Ar–H and 3thienyl Hs); 13C NMR (100 MHz, DMSO)d
ppm: 55.65 (C-3), 56.38 (OCH3), 59.42 (2 OCH3), 60.39
(OCH3), 64.00 (C-4), 104.33 (2C), 114.91 (2C), 119.05
(2C), 126.09, 126.64, 127.83, 130.95, 133.11, 136.96,
138.03, 153.85 (2C), 156.31 (ArC), 164.43 (C=O); EIMS
m/z (%rel. abundance): 425.03 (M?, 11.38); Anal. Calcd
for C23H23NO5S: C 64.92, H 5.45, N 3.29. Found: C 65.08,
H 5.53, N 3.41.
(s, 3H, OCH3), 3.65 (s, 6H, 2OCH3), 4.47 (d, 1H, H-4),
5.60 (d, 1H, H-3), 6.51 (s, 2H, Ar–H), 7.26–7.33 (m, 3H,
Ar–H), 7.36–7.43 (m, 5H, Ar–H), 7.66 (d, 1H, Ar–H,
J = 7.8 Hz); 13C NMR (100 MHz, DMSO)d ppm: 55.71
(C-3), 60.05 (C-4), 61.35 (2 OCH3), 63.41 (OCH3), 94.93
(2C), 122.316, 127.66 (2C), 127.74 (2C), 128.43 (2C),
128.77 (2C), 130.25, 132.84, 133.09, 134.79, 136.165,
153.28 (2C) (ArC), 165.008 (C=O); EIMS m/z (%rel.
abundance): 466.86 (M?, 4.06), 468.97 (M?2, 5.07); Anal.
Calcd for C24H22BrNO4: C 61.55, H 4.73, N 2.99. Found:
C 61.70, H 4.82, N 3.08.
4-(2-Bromophenyl)-3-chloro-1-(3,4,5-trimethoxyphenyl)aze-
tidin-2-one (8h) Was prepared from N-(2-bromobenzyli-
dene)-3,4,5-trimethoxybenzenamine (6f) and acyl chloride
7. Yield 42 %, grayish brown crystals, mp 118–119 °C. IR
(KBr, t cm-1): 3016 (CH aromatic), 2995, 2981 (CH ali-
phatic), 1762 (C=O); 1H NMR (400 MHz, DMSO-d6)d
ppm: 3.61 (s, 3H, OCH3), 3.67 (s, 6H, 2OCH3), 5.31 (d,
1H, H-4, J = 1.6 Hz), 5.63 (d, 1H, H-3, J = 1.6 Hz), 6.57
(s, 2H, Ar–H), 7.32–7.43 (m, 3H, Ar–H), 7.76 (d, 1H, Ar–
H, J = 9 Hz); 13C NMR (100 MHz, DMSO)d ppm: 56.32
(C-4), 60.57 (2 OCH3), 62.28 (OCH3) 64.51 (C-3), 96.02
(2C), 122.89, 128.82, 131.34 (2C), 132.67, 133.75, 134.32,
135.24, 153.79 (2C) (ArC), 160.86 (C=O); EIMS m/z
(%rel. abundance): 424.84 (M?, 15.83), 425.87 (M?H,
3.09), 426.85 (M?2, 20.42), 427.86 (M?3, 3.95), 428.84
(M?4, 5.52); Anal. Calcd for C18H17BrClNO4: C 50.67, H
4.02, N 3.28. Found: C 50.79, H 4.08, N 3.41.
4-(2-Bromophenyl)-1-(3,4,5-trimethoxyphenyl)-3-(2-thienyl)aze-
tidin-2-one (8k) Was prepared from N-(2-bromobenzyli-
dene)-3,4,5-trimethoxybenzenamine (6f) and acyl chloride
7. Yield 61 %, buff crystals, mp 109–110 °C. IR (KBr, t
cm-1): 3066 (CH aromatic), 2951, 2825 (CH aliphatic),
1
1764 (C=O); H NMR (400 MHz, DMSO-d6)d ppm: 3.60
(s, 3H, OCH3), 3.66 (s, 6H, 2 OCH3), 4.83 (d, 1H, H-4,
J = 2.4 Hz), 5.58 (d, 1H, H-3, J = 2.4 Hz), 6.56 (s, 2H,
Ar–H), 7.07–7.56 (m, 6H, 3Ar–H and 3H thienyl H), 7.72
(d, 1H, Ar–H); 13C NMR (100 MHz, DMSO)d ppm: 56.25
(C-3), 58.96 (C-4), 60.58 (2 OCH3), 62.96 (OCH3), 95.43
(2C), 122.93, 126.49, 127.15, 127.79, 129.02, 130.99 (2C),
133.24, 133.66, 134.76, 136.01, 136.68, 153.81 (2C) (ArC),
164.50 (C=O); EIMS m/z (%rel. abundance): 472.82 (M?,
33.19), 474.83 (M?2, 34.90); Anal. Calcd for C22H20-
BrNO4S: C 55.70, H 4.25, N 2.95. Found: C 55.84, H 4.31,
N 3.03.
4-(2-Bromophenyl)-3-chloro-1-(3,4,5-trimethoxyphenyl)-3-
methylazetidin-2-one (8i) Was prepared from N-(2-bro-
mobenzylidene)-3,4,5-trimethoxybenzenamine (6f) and
acyl chloride 7. Yield 77 %, buff crystals, mp 150–151 °C.
IR (KBr, t cm-1): 3001 (CH aromatic), 2927, 2889 (CH
4-(3-Bromophenyl)-1-(3,4,5-trimethoxyphenyl)-3-(2-thienyl)aze-
tidin-2-one (8l) Was prepared from N-(3-bromobenzyli-
dene)-3,4,5-trimethoxybenzenamine (6g) and acyl chloride
7. Yield 30 % (65 % by microwave method), buff crystals,
mp 118–119 °C. IR (KBr, t cm-1): 3001 (CH aromatic),
2958, 2924 (CH aliphatic), 1751; (C=O) 1H NMR
(400 MHz, DMSO-d6)d ppm: 3.59 (s, 3H, OCH3), 3.66 (s,
6H, 2OCH3), 4.87(d, 1H, H-3, J = 2.3 Hz), 5.36 (d, 1H,
H-4, J = 2.3 Hz), 6.61 (s, 2H, Ar–H), 7.06-7.08 (m, 1H,
thienyl-H), 7.16 (d, 1H, thienyl-H), 7.36–7.40 (t, 1H,
thienyl-H, J = 7.84 Hz), 7.53–7.58 (m, 3H, Ar–H), 7.80 (s,
1H, Ar–H); 13C NMR (100 MHz, DMSO)d ppm: 56.23 (C-
3), 58.96 (2 OCH3), 60.57 (OCH3), 62.78 (C-4) 95.62 (2C),
122.72, 126.29, 126.56, 126.84, 127.86, 130.09, 131.67,
132.13, 133.18, 134.68, 136.41, 140.21, 153.71 (2C) (ArC),
164.53 (C=O); EIMS m/z (%rel. abundance): 472.90 (M?,
7.63), 474.91 (M?2, 8.90); Anal. Calcd for C22H20-
BrNO4S: C 55.70, H 4.25, N 2.95. Found: C 55.89, H 4.31,
N 3.07.
1
aliphatic), 1757 (C=O); H NMR (300 MHz, DMSO-d6)d
ppm: 2.12 (s, 3H, CH3), 3.63 (s, 3H, OCH3), 3.67 (s, 6H,
2OCH3), 5.81 (s, 1H, H-4), 6.64 (s, 2H, Ar–H), 7.06–7.39
(m, 3H, Ar–H), 7.73–7.81 (d, 1H, Ar–H); 13C NMR
(100 MHz, DMSO)d ppm: 24.88 (CH3) 56.43 (2 OCH3),
60.60 (OCH3) 67.29 (C-3), 69.04 (C-4), 96.36 (2C),
123.12, 128.16, 128.50, 130.78, 131.04, 132.77, 133.77,
135.39, 153.76 (2C) (ArC), 163.85 (C=O); EIMS m/z
(%rel. abundance): 438.95 (M?, 1.48), 440.97 (M?2,
2.12); Anal. Calcd for C19H19BrClNO4: C 51.78, H 4.35, N
3.18. Found: C 51.87, H 4.42, N 3.25.
4-(2-Bromophenyl)-1-(3,4,5-trimethoxyphenyl)-3-phenylaze-
tidin-2-one (8j) Was prepared from N-(2-bromobenzyli-
dene)-3,4,5-trimethoxybenzenamine (6f) and acyl chloride
123

Synthesis and cytotoxic activity of certain trisubstituted azetidin-2-one derivatives…
4-(4-Bromophenyl)-1-(3,4,5-trimethoxyphenyl)-3-phenylaze-
tidin-2-one (8m) Was prepared from N-(4-bromoben-
zylidene)-3,4,5-trimethoxybenzenamine (6h) and acyl
chloride 7. Yield 19 % (45 % by microwave method),
yellow crystals, mp 113–114 °C. IR (KBr, t cm-1): 3001
(CH aromatic), 2954, 2850 (CH aliphatic), 1741 (C=O); 1H
NMR (300 MHz, DMSO-d6)d ppm: 3.59 (s, 3H, OCH3),
3.65 (s, 6H, 2OCH3), 4.47(d, 1H, H-4), 5.34 (d, 1H, H-3),
6.61 (s, 2H, Ar–H), 7.38–7.41 (m, 5H, Ar–H), 7.47 (d, 2H,
Ar–H, J = 7.8 Hz), 7.60 (d, 2H, Ar–H, J = 7.8 Hz); 13C
NMR (100 MHz, DMSO)d ppm: 55.75 (C-3), 60.04
(2OCH3), 61.32 (OCH3), 63.43 (C-4), 95.16 (2C), 121.64,
127.50, 127.73 (2C), 128.83 (2C), 128.87 (2C), 131.89
(2C), 132.78, 134.12, 134.60, 136.99, 153.18 (2C) (ArC),
165.015 (C=O); EIMS m/z (%rel. abundance): 468.15
(M?, 73.91), 469.15 (M?H, 13.59). Anal. Calcd for C24-
H22BrNO4: C 61.55, H 4.73, N 2.99. Found: C 61.72, H
4.80, N 3.03.
425.14 (M?2, 0.86); Anal. Calcd for C24H22ClNO4: C 68.00,
H 5.23, N 3.30. Found: C 68.14, H 5.31, N 3.43.
4-(4-Chlorophenyl)-1-(3,4,5-trimethoxyphenyl)-3-(2-thienyl)aze-
tidin-2-one (8p) Was prepared from N-(4-chlorobenzyli-
dene)-3,4,5-trimethoxybenzenamine (6i) and acyl chloride
7. Yield 89 %, buff crystals, mp 152–153 °C. IR (KBr, t
cm-1): 3111 (CH aromatic), 2954, 2825 (CH aliphatic),
1
1761 (C=O); H NMR (300 MHz, DMSO-d6)d ppm: 3.58
(s, 3H, OCH3), 3.65 (s, 6H, 2OCH3), 4.78 (d, 1H, H-4,
J = 2.4 Hz), 5.35 (d, 1H, H-3, J = 2.4 Hz), 6.58 (s, 2H,
Ar–H), 7.06–7.15 (m, 3H, thienyl), 7.47 (d, 2H, Ar-H,
J = 8.4 Hz), 7.55 (d, 2H, Ar-H, J = 8.4 Hz); 13C NMR
(100 MHz, DMSO)d ppm: 56.25 (C-3), 59.12 (2OCH3),
60.55 (OCH3), 62.85 (C-4), 95.65 (2C), 126.54, 126.78,
127.85, 129.09 (2C), 129.50 (2C), 133.21, 133.77, 134.66,
136.50, 136.53, 153.70 (2C) (ArC), 164.51 (C=O); EIMS
m/z (%rel. abundance): 429.15 (M?, 17.50), 431.10 (M?2,
6.79). Anal. Calcd for C22H20ClNO4S: C 61.46, H 4.69, N
3.26. Found: C 61.62, H 4.73, N 3.42.
4-(4-Bromophenyl)-1-(3,4,5-trimethoxyphenyl)-3-(2-thienyl)aze-
tidin-2-one (8n) Was prepared from N-(4-bromobenzyli-
dene)-3,4,5-trimethoxybenzenamine (6h) and acyl chloride
7. Yield 39 % (56 % by microwave method), pale brown
crystals, mp 173–174 °C. IR (KBr, t cm-1): 3111 (CH
aromatic), 2954, 2823 (CH aliphatic), 1759 (C=O); 1H
NMR (300 MHz, DMSO-d6)d ppm: 3.58 (s, 3H, OCH3),
3.65 (s, 6H, 2OCH3), 4.77 (d, 1H, H-4, J = 2.4 Hz), 5.34
(d, 1H, H-3, J = 2.4 Hz), 6.59 (s, 2H, Ar–H), 7.06–7.15
(m, 3H, thienyl Hs), 7.48 (d, 2H, Ar–H, J = 8.4 Hz), 7.61
(d, 2H, Ar–H, J = 8.4 Hz); 13C NMR (100 MHz, DMSO)d
ppm: 56.26 (C-3), 59.08 (2 OCH3), 60.56 (OCH3), 62.90
(C-4), 95.65 (2C), 122.35, 126.55, 126.79, 127.85, 129.37
(2C), 132.42 (2C), 133.20, 134.67, 136.52, 136.93, 153.71
(2C) (ArC), 164.51 (C=O); EIMS m/z (%rel. abundance):
472.88 (M?, 26.64), 474.88 (M?2, 27.97); Anal. Calcd for
C22H20BrNO4S: C 55.70, H 4.25, N 2.95. Found: C 55.89,
H 4.28, N 3.01.
Biological study
Measurement of cytotoxicity by SRB assay
The cytotoxic activity of the newly synthesized compounds
was measured in vitro using SRB assay according to the
previously reported method (Skehan et al. 1990). Cells
were plated in 96-multiwell microtiter plates (104 cells/
well) for 24 h before treatment with the compound(s) to
allow attachment of cells to the walls of the plates. Test
compounds were dissolved in DMSO and diluted with
saline to the appropriate volume. Different concentrations
of the test compounds (0, 5, 12, 25, and 50 lg/mL) were
added to the cell monolayer. Triplicate wells were prepared
for each individual dose. Monolayer cells were incubated
with the compound(s) for 48 h at 37 °C in an atmosphere
of 5 % CO2. After 48 h, cells were fixed, washed and
stained for 30 min with 0.4 % (wt/vol) with SRB dissolved
in 1 % acetic acid. Unbound dye was removed by four
washes with 1 % acetic acid, and attached stain was
recovered with 2 Tris EDTA buffer. Color intensity was
measured in an ELISA reader. The relationship between
surviving fraction and drug concentration was plotted to
get the survival curve. The concentration required for 50 %
inhibition of cell viability (IC50) was calculated. The
results are given in Tables 1 and 3 and graphically pre-
sented in Fig. 1.
4-(4-Chlorophenyl)-1-(3,4,5-trimethoxyphenyl)-3-pheny-
lazetidin-2-one (8o) Was prepared from N-(4-chloroben-
zylidene)-3,4,5-trimethoxybenzenamine (6i) and acyl
chloride 7. Yield 22 % (49 % by microwave method), yellow
crystals, mp 97–98 °C. IR (KBr, t cm-1): 3030 (CH aro-
1
matic), 2997, 2933(CH aliphatic), 1743 (C=O); H NMR
(300 MHz, DMSO-d6)d ppm: 3.59 (s, 3H, OCH3), 3.65 (s,
6H, 2OCH3), 4.48 (d, 1H, H-4, J = 2.1 Hz), 5.36 (d, 1H,
H-3, J = 2.1 Hz), 6.61 (s, 2H, Ar–H), 7.36–7.42 (m, 5H, Ar–
H), 7.47 (d, 2H, Ar–H, J = 8.4 Hz), 7.55 (d, 2H, Ar–H,
J = 8.4 Hz); 13C NMR (100 MHz, DMSO)d ppm: 56.25 (C-
3), 60.56 (2 OCH3), 61.81 (OCH3), 63.90 (C-4), 95.60 (2C),
122.16, 128.03 (2C), 129.38 (2C), 129.40 (4C), 132.41,
133.28, 134.55, 135.12, 137.49, 153.68 (2C) (ArC), 165.56
(C=O); EIMS m/z (%rel. abundance): 423.12 (M?, 0.89),
In vitro tubulin polymerization assay
MCF-7 and HCT-116 cells were obtained from American
Type Culture Collection, and cultured using DMEM
123

S. Elmeligie et al.
was performed according to the manufacturer’s
instructions.
Table 1 In vitro cytotoxic activitiy of the tested compounds 8a–p on
MCF-7 and HCT-116 cell lines
Compound no.
MCF-7 (IC50)a (lM)
HCT (IC50)a (lM)
Molecular modeling study
8a
11.18
21.80
9.27
11.67
22.67
28.09
41.64
16.84
22.97
17.46
8.62
8b
The PDB (code: 1SA0) was chosen as the template for the
modeling study of compounds 8n and 8o bound to
colchicines-binding site. The crystal structure was obtained
from RCSB protein data bank (http://www.rcsb.org/pdb/
home/home.do). The molecular docking study was carried
out using CDOCKER protocol within Discovery Studio
version 2.55. For protein preparation, only chain A and B
with co crystallized Colchicine 1 was retained, the hydro-
gen atoms were added and optimized, the water molecules
and impurities were removed. For ligand preparation, the
3D structures of the tested compounds were built using
ChemBioDraw Ultra 11.0 and minimized using discovery
studio 2.55. The validation results showed the same bind-
ing interactions of the co-crystallized and the re-docked
8c
8d
20.82
10.81
36.01
8.22
8e
8f
8g
8h
9.37
8i
18.15
7.25
10.39
6.25
8j
8k
28.45
156.2
8.54
8.70
8l
185.7
20.02
7.44
8m
8n
6.74
8o
7.07
8.68
˚
ligand with rmsd of 0.95 A The docking study was per-
8p
10.93
12.35
9.25
formed by inserting the tested compounds into the
colchicines-binding site. The type of interactions between
the compounds and protein were analyzed.
Colchicine
3.8
a
IC50 dose of the compound that inhibits tumor cell proliferation by
50 %
Results and discussion
Chemistry
(Invitrogen/Life Technologies) supplemented with 10 %
FBS (Hyclone), 10 ug/mL of insulin (Sigma-Aldrich, MO,
USA), and 1 % penicillin–streptomycin. All of the other
chemicals and reagents were purchased from Sigma, or
Invitrogen. Plate cells (cells density 1.2–1.8 9 10,000
cells/well) in a volume of 100 lL complete growth med-
ium ?100 ul of the tested compound with the IC50 conc.
per well in a 96-well plate for 24 h before the enzyme
assay for tubulin.
The preparation of target combretastatin-based analogues
8a–p was based upon incorporation of 4-methoxyphenyl or
3,4,5-trimethoxyphenyl scaffold at N-1 position of the b-
lactam ring (mimicking ring A of CA-4) and a substituted
phenyl ring at C-4 of the b-lactam ring (mimicking ring B
of CA-4), (Fig. 1). As shown in Scheme 1, the synthesis
firstly involves the formation of the imine precursors 6a–
i by condensation of the appropriately substituted primary
aromatic amines 4 and certain substituted benzaldehydes 5
Tubulin polymerization inhibitory activity was deter-
mined using human type b-tubulin SEB870HU assay kit
(Cloud-Clone Corp. USA). The procedure of the used kit
Fig. 1 Some tubulin binding agents and the newly synthesized compounds 8a–p
123

Synthesis and cytotoxic activity of certain trisubstituted azetidin-2-one derivatives…
to give the desired imines in high decent yields. The target
b-lactam derivatives 8a–p were obtained adopting the
Staudinger cycloaddition reaction, which initiated by the
nucleophilic attack of the imine derivatives 6a–i to an
insitu generated ketene (from an appropriate substituted
acetyl chloride 7 and TEA) following previously reported
procedure (Banik et al. 2005) (Scheme 1). Stereochemistry
of 8a–p depends upon several factors; reaction conditions,
order of addition of the reagents and substituents present on
both the acid chloride and the imine (Xu 2009). In this
work, the target compounds 8a–p showed trans arrange-
ment for H-3 and H-4 as evidenced by a pair of coupled
doublets with coupling constants about, J = 2 Hz. No cis
isomers were isolated in this series, likely due to steric
hindrance between the substituents in 3- and 4-positions of
the azetidin-2-one ring. Some b-lactam derivatives, namely
8l–o, were synthesized adopting microwave irradiation by
applying Microwave-induced Organic Reaction Enhance-
ment (MORE) chemistry technique, starting with the
appropriate imine precursors 6g–i and certain substituted
acetyl chlorides 7 in chlorobenzene and TEA for 10 min
(Bose et al. 1995; Bandyopadhyay and Banik 2010)
(Scheme 2). The b-lactam derivatives prepared by micro-
wave irradiation technique were isolated in high yields
with stereochemistry similar to that obtained by traditional
methods.
Scheme 1 Reagents and conditions: a ethanol, reflux, 5–12 h; b CH2Cl2, TEA, reflux, 7–12 h
123

S. Elmeligie et al.
Biological result
compound 8i containing both chloro and methyl groups at
C-3 of the ring. Compound 8k with 2-thienyl substituent at
C-3 showed the weakest activity among this series of
compounds against MCF-7, but relatively good activity
against HCT-116 cell line (IC50 = 8.70 lM). Among the
series of compounds 8k, 8l, 8n, 8p, with 3,4,5-trimethox-
yphenyl at N-1 and 2-thienyl substituent at C-3, it was
obvious that the position of the substituents on the phenyl
ring at C-4 of the azetidin-2-one ring is critical on the
cytotoxicity of these compounds. It was observed that
compounds with bromo or chloro substituents at ortho or
para position of the phenyl ring displayed excellent to
moderate cytotoxic activity with IC50 range of
6.74–28.45 lM against both MCF-7 and HCT-116 cell
lines. On the other hand, compound 8l with bromo sub-
stituent at meta position showed dramatic reduction in
cytotoxicity against both tested human cancer cell lines.
In conclusion, the most active azetidin-2-one derivatives
contain at C-4 of the ring either 3,4,5-trimethoxyphenyl
substitution or 2- or 4-bromo or chloro substituted phenyl
ring. Moreover, compounds with 3,4,5-trimethoxy phenyl
substituents at N-1 showed generally better cytotoxic
activity compared with the compounds having 4-methox-
yphenyl substituent. On the other hand, compounds which
are disubstituted at C-3 were found to display weaker
cytotoxic activity compared with their monosubstituted
analogues.
Cytotoxic activity
The newly synthesized b-lactam derivatives 8a–p were
screened for their cytotoxic activity against human breast
cancer MCF-7 and the human colon cancer HCT-116 cell
lines, using sulphorhodamine B (SRB) stain colorimetric
assay. The drug concentration required to inhibit the cell
growth by 50 % (IC50) was determined and the results were
displayed and represented graphically (Table 1; Fig. 2).
For comparison, the IC50 of the standard compound,
colchicine, was evaluated. The results revealed that all
tested compounds displayed cytotoxic activity against both
tested cell lines. Compounds 8a, 8c, 8e, 8g, 8h, 8j, 8m, 8n,
8o and 8p showed cytotoxic activity against MCF-7 cell
line superior to colchicine with IC50 range 6.74-11.18 lM,
while compounds 8b, 8d, 8i and 8k demonstrated moderate
activity. On the other hand, weak cytotoxic effect was
observed by the compounds 8f and 8l. Further, the b-lac-
tam derivatives 8a, 8h, 8i, 8j, 8k, 8n, 8o and 8p were found
to be the most potent against HCT-116 cell line with IC50
range 6.25–11.67 lM.
Based on the cytotoxicity data, the structure–activity
relationship (SAR) for these trisubstituted azetidin-2-one
derivatives has been elucidated. Keeping constant the
3,4,5-trimethoxyphenyl ring at N-1 and 2-bromophenyl
substituent at C-4, the effect of modifications on the
position 3 of the azetidin-2-one ring has been examined in
compounds 8h, 8i, 8j and 8k. Compound 8j having a
phenyl substituent exhibited very good anticancer efficacy
towards both MCF-7 and HCT-116 cell lines (IC50 = 7.25
and 6.25 lM) respectively. Slight reduction in cytotoxic
activity was observed when the phenyl group was replaced
by chloro substituent (8h). Further decrease in the activity
against MCF-7 and HCT-116 cell lines was observed in the
Tubulin polymerization inhibition assay
The most potent compounds in SRB assay, 8g, 8j, 8n and
8o, were further investigated for inhibition of tubulin
polymerization aganist MCF-7 and HCT-116 cancer cell
lines by applying enzyme linked immunosorbent assay
(ELISA) using human b-tubulin assay kit SEB870Hu
obtained from Cloud-Clone Corp. USA. The inhibitory
Scheme2 Reagents and
conditions: a chlorobenzene,
TEA, microwave, 10 min
123

Synthesis and cytotoxic activity of certain trisubstituted azetidin-2-one derivatives…
Fig. 2 Cytotoxic activity of the tested compounds 8a–p on MCF-7 and HCT-116 cell lines
Table 2 Percentage inhibition of tubulin polymerization on human MCF-7 and HCT-116 cell lines at IC50 for the compounds 8g, 8j, 8n and 8o
Compound
no.
IC50 (lM)
MCF-7
% inhibition of tubulin
polymerization (MCF-7)
IC50 (lM)
HCT-116
% inhibition of tubulin
polymerization (HCT-116)
8g
8.22
7.25
85.18
81.12
79.10
78.11
86.90
17.46
6.25
7.44
8.68
3.8
83.63
85.60
86.86
84.83
84.41
8j
8n
6.74
8o
7.07
Colchicine
12.35
Molecular modeling studies
activity of tubulin was given as the percentage inhibition at
IC50 concentration of each compound (Table 2). The
results showed that all tested compounds produced signif-
icant tubulin suppression compared to the reference
antimitotic agent colchicine. Therefore, cytotoxic activity
of the tested compounds may be referred to their anti-
tubulin activity.
Molecular modeling simulation study was performed to
investigate the probable binding modes of the most active
Table 3 In vitro cytotoxic activity of the tested compounds 8g, 8j, 8n
and 8o on Hs371.T and CCD18co cell lines
Compound no.
IC50 (lM)a
Evaluation of toxicity in normal cells
Hs371.T
CCD18co
8g
88.8
38.13
15.88
30.44
29.57
58.12
Equally important, compounds 8g, 8j, 8n and 8o were
further investigated for toxicity on normal breast Hs371.T
cell line and normal colon CCD18co cell line using
sulphorhodamine B (SRB) stain colorimetric assay. Results
in Table 3 revealed that compounds 8g and 8o displayed
the lowest toxicity on normal Hs371.T in comparison to
colchicines as the reference compound.
8j
43.23
44.03
77.38
67.21
8n
8o
Colchicine
a
IC50 dose of the compound that inhibits tumor cell proliferation by
50 %
123

S. Elmeligie et al.
Fig. 3 2D and 3D interaction of compound 8n in the colchicine binding site of tubulin (PDB code 1SA0); showing key hydrogen bonding
interactions (dotted lines)
Fig. 4 2D and 3D interaction of compound 8o in the colchicine binding site of tubulin (PDB code 1SA0); showing key hydrogen bonding
interactions (dotted lines)
compounds, 8n and 8o, using the reported X-ray structure
of tubulin co-crystallized with N-deacetyl-N-(2-mercapto
acetyl) colchicine (DAMA colchicine, PDB code: 1SA0)
(Wei et al. 2004). The colchicine-binding site was mainly
buried in the b-subunit of tubulin, in addition to some
limited interaction with the a-subunit, while surrounded by
H7 and H8 a-helices, S8 and S9 b-strands and T7 loop,
which interact with the colchicine-site ligand (Dorleans
et al. 2009). Researches have proved that the most
important residues for colchicine-binding are Val318 and
123

Synthesis and cytotoxic activity of certain trisubstituted azetidin-2-one derivatives…
the binding to tubulin and microtubule inhibition. Biochemistry
37:8356–8368
Cys241. Methoxy group in ring A of colchicine can be
cross linked with Cys241 (Andreu et al. 1998). Also
Thr179, Lys 352, Val 181, Leu248 and Ala250 have been
reported to play a crucial role in the interaction of azetidin-
2-ones with the colchicines-binding site (O’Boyle et al.
2011b). Results obtained from docking imitation were
presented in Figs. 3 and 4. Hydrogen bonding interaction
between Cys 241 and methoxy group of the compounds 8n
and 8o was found to be comparable to colchicine. These
binding parallels may rationalize the cytotoxic potency
observed by these compounds. Also, hydrogen bonding
interaction between oxygen at C-2 and Asp 251, Leu 252,
Leu 255 may rationalize the increased activity of these
compounds. In addition, compound 8n showed hydrogen
bonding interaction between thiophene ring sulfur and Ala
250, (Fig. 3). Moreover, strong p–p bond stabilizes the
interaction between Lys 254 amino acid residue and the
aromatic ring system at C-3 in these compounds.
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Synthesis and evaluation of azetidinone analogues of combre-
In this research, new series of azetidin-2-one derivatives
8a–p were synthesized with the aim to rigidify the tradi-
tionally isomerizable double bond in CA-4 that suppresses
its activity. The synthesized compounds were evaluated for
their cytotoxic activity against MCF-7 and HCT-116 can-
cer cell lines. It was obvious that, presence of 3,4,5-tri-
methoxy phenyl scaffold at N1 or C-4 and a 2-thienyl or
phenyl group at C-3 was found to increase the cytotoxic
activity of these compounds. Four compounds 8g, 8j, 8n
and 8o showed higher percentage suppression of tubulin
(79.09, 78.11, 81.12 and 85.18) respectively in comparison
with colchicine against MCF-7 cell line and (86.86, 86.83,
85.63 and 83.63) respectively against HCT-116 cell line.
Molecular modeling studies were performed to rationalize
the observed cytotoxic activity by the synthesized com-
pounds. These compounds could be considered a promising
leads for further investigation.
tastatin A-4 as tubulin targeting agents.
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DG, Zisterer DM, Meegan MJ (2011b) Synthesis, evaluation and
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Acknowledgments The authors would like to thank all members of
the department of Cancer Biology, National Cancer Institute, Cairo,
Egypt, for carrying out the cytotoxicity testing and Dr. Essam
Rashwan, Head of Confirmatory Diagnostic Unit, Vacsera, Egypt for
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