Synthesis of benzimidazoles bearing oxadiazole nucleus as anticancer agents

Article abstract of DOI:10.1016/j.ejmech.2012.04.027

Starting from 4-(1H-benzo[d]imidazol-2-yl)-4-oxobutanehydrazide (1), twenty new 1-(1H-benzo[d]imidazol-2-yl)-3-(1,3,4-oxadiazol-5-substituted derivatives-2-yl)propan-1-ones (1b-A_7-26) were synthesized under microwave irradiation in good yields. Further, compound 1b-A₇ was reacted with different secondary amines under microwave irradiation to produce five novel 1-(1H-benzo[d]imidazol-2-yl)-3-(5-(methyl substituted)-1,3,4-oxadiazol-2-yl)propan-1-ones (1b-A_{7a -e}). The title compounds were screened for their in vitro anticancer activity at National Cancer Institute (NCI), USA; at a single dose (10 μM) in NCI 60 cell line panel and results showed significant to good anticancer activity. One compound, 1b-A₁₈ (NSC: 759205), 1-(1H-benzo[d]imidazol-2-yl)-3-(5-(2,4-dichlorophenyl)-1,3,4-oxadiazol-2-yl) propan-1-one, emerged as lead compound; it was selected for five-dose level screening and found to have significant growth inhibition activity.

Full text of DOI:10.1016/j.ejmech.2012.04.027

European Journal of Medicinal Chemistry 54 (2012) 855e866
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
Synthesis of benzimidazoles bearing oxadiazole nucleus as anticancer agents
*
Mohd Rashid, Asif Husain , Ravinesh Mishra
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Jamia Hamdard (Hamdard University), Hamdard Nagar, New Delhi 110062, India
h i g h l i g h t s
g r a p h i c a l a b s t r a c t
< 25 Novel benzimidazoles bearing
oxadiazole nucleus were synthe-
sized under microwave irradiation.
< Initial screening of compounds
showed significant to good in vitro
anticancer activity.
< One compound, 1beA18, emerged as
lead compound and screened at five
dose level.
< The compound 1beA18 can be used
as a template for designing potential
anticancer agents.
a r t i c l e i n f o
a b s t r a c t
Article history:
Starting from 4-(1H-benzo[d]imidazol-2-yl)-4-oxobutanehydrazide (1), twenty new 1-(1H-benzo[d]
Received 16 November 2011
Received in revised form
15 April 2012
Accepted 18 April 2012
Available online 9 May 2012
imidazol-2-yl)-3-(1,3,4-oxadiazol-5-substituted derivatives-2-yl)propan-1-ones (1beA7e26
)
were
synthesized under microwave irradiation in good yields. Further, compound 1beA7 was reacted with
different secondary amines under microwave irradiation to produce five novel 1-(1H-benzo[d]imidazol-
2-yl)-3-(5-(methyl substituted)-1,3,4-oxadiazol-2-yl)propan-1-ones (1beA7aee). The title compounds
were screened for their in vitro anticancer activity at National Cancer Institute (NCI), USA; at a single dose
(10
mM) in NCI 60 cell line panel and results showed significant to good anticancer activity. One
Keywords:
compound, 1beA18 (NSC: 759205), 1-(1H-benzo[d]imidazol-2-yl)-3-(5-(2,4-dichlorophenyl)-1,3,4-
oxadiazol-2-yl)propan-1-one, emerged as lead compound; it was selected for five-dose level screening
and found to have significant growth inhibition activity.
Benzimidazole
1,3,4-oxadiazole
Microwave-assisted
Anticancer activity
Ó 2012 Elsevier Masson SAS. All rights reserved.
1. Introduction
therapeutic index, with significant toxicity from effective drug
doses or tumor recurrence at low drug doses. Searching of new
Cancer is a collection of different life threatening diseases
characterized by uncontrolled growth of cells, leading to invasion of
surrounding tissue and often spreading to other parts of the body.
When it comes to understanding and controlling cancer, scientists
are now working from a position of strength e a sturdy foundation
of knowledge about cancer built over the past 50 years. There is an
urgent need for novel effective drug regimens for the treatment of
cancer because the current chemotherapy suffers from a slim
anticancer agents having heterocyclic nucleus continues world-
wide at various laboratories [1e3].
Scientists from worldwide have reported remarkable antitumor/
antiproliferative/anticancer [4e13], anti-inflammatory [12], anti-
viral including anti-HIV [13,14], antibacterial [15e17], antifungal
[18,19] and antioxidant [19,20] activities of different benzimidazole
derivatives. Similarly, 1,3,4-oxadiazole is a class of heterocyclic
compound that have attracted significant interest in medicinal
chemistry as they have a wide range of pharmaceutical and phar-
macological applications including potential antitumor, anti-
proliferative or anticancer activities [21e26]. Benzimidazole
clubbed with other heterocyclic moieties including the oxadiazole
* Corresponding author. Tel.: þ91 11 26059681/688x5890; fax: þ91 11
16988874; þ91 9891116086 (mobile).
E-mail address: drasifhusain@yahoo.com (A. Husain).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2012.04.027

856
M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
nucleus resulted in compounds with improved biological activities
including anticancer activity [10e13,27]. Motivated by the these
observations, it was thought worthwhile to prepare new hybrid
compounds that comprise both benzimidazole and 1,3,4-oxadiazole
ring systems with a view to obtaining promising anticancer agents.
Therefore, several benzimidazoles having 1,3,4-oxadiazole moie-
ties were synthesized and screened for their in vitro anticancer
activities at National Cancer Institute (NCI), USA. The selected
compounds were submitted to NCI and granted NCS codes as NCS:
755135 (1beA7), NCS: 755139 (1beA10), 759200 (IbeA11), NCS:
759201 (IbeA12), NCS: 759202 (IbeA13), NCS: 759203 (IbeA14), NCS:
759204 (IbeA15), NCS: 759205 (IbeA18), NCS: 759206 (IbeA23), NCS:
759207 (IbeA25), NCS: 759208 (IbeA26), NCS: 759209 (IbeA7a), NCS:
759210 (IbeA7b), NCS: 759211 (IbeA7c), NCS: 760445 (IbeA7d) and
NCS: 759212 (IbeA7e) for screening their anticancer activity against
exhibit significant growth inhibition at a single dose of 10 mM are
evaluated against the 60 cell panel at five concentration levels.
4. Results and discussion
4.1. Chemistry
Overall twenty-eight compounds were prepared as outlined in
Figs.1, 2. The structures assigned to the compounds were supported
by the results of elemental analysis as well as IR, 1H NMR, 13C NMR
and Mass spectral data results. In general, IR value showed peaks at
around 3424 (NeH, NH2), 3350 (NeH, NH), 3020(CeH, AreH,
aromatic hydrogen), 2950, 2860 (CeH, CH2), 1720 (C]O), 1674
(C]N), 1562, 1504, 1485 (C]C, ring stretch), 1308 (NeN]C), 1164
(CeOeC, asymmetric), 1028 (CeOeC, symmetric), 834 (CeN,
aliphatic stretch). 1H NMR spectra peak of the respective protons
of the synthesized compounds verified on the basis of their
full NCI 60 cell line panel at single dose (10 mM), among these, the
compound IbeA18 (NCS: 759205) exhibited significant growth inhi-
bition at a single dose and further selected for anticancer screening at
five dose concentration level against full NCI 60 cell panel.
chemical shifts (
compounds showed two triplets at around
could be accounted for two methylene groups (eCH2eCH2e),
doublet, triplet at around 7.6, 7.5 and 7.3 indicative of benzimid-
azole hydrogen and a singlet at around 11.9 indicative of ring HeN
(D2O exchangeable). Other peaks were observed at appropriate
values supporting the structure. 13C NMR spectra peaks at
175.5
for carbonyl group, 162.7, 159.6 indicative of oxadiazole carbon,
154.96 for C]N and 138.27, 132.21, 130.19, 128.57, 124.65, 123.64
indicative of AreC (aromatic carbon), 38.7, 27.4 accounted for two
d
), multiplicities, and coupling constants (J). All the
d
2.8 and 3.3 which
2. Chemistry
d
d
The
oxobutanoic acid (1), was synthesized by reacting o-phenylene
diamine with -ketoglutaric acid in presence of HCl. Compound (1)
starting
material,
4-(1H-benzo[d]imidazol-2-yl)-4-
d
d
a
d
was treated with hydrazine hydrate through ester intermediate to
furnish 4-(1H-benzo[d]imidazol-2-yl)-4-oxobutanehydrazide (Ib).
Different types of acids were reacted with compound (Ib) under
microwave irradiation to get 1-(1H-benzo[d]imidazol-2-yl)
d
d
d
methylene groups (eCH2eCH2e). The mass spectra (ESI-MS)
showed the presence of peak at definite m/z value in accordance to
the molecular formula. In case of aryl groups having chloro-
substituent(s), the molecular ion peak appeared as cluster of
peaks. The elemental analysis results were found to be within
ꢀ0.4% deviation from the theoretical values.
-3-(1,3,4-oxadiazol-5-substituted-2-yl)propan-1-ones (IbeA7e26
)
(Fig. 1). The purity of compounds was checked by single-spot TLC
using toluene: ethyl acetate: formic acid (5:4:1) and benzene:
acetone (9:1) solvent systems and spots located under iodine vapors/
UV light. The final products were purified by flash chromatography
column (hexane/ethyl acetate gradient, 8:2, gradient varied
compound to compound). The compound IbeA7 was further reacted
with different types of secondary amines in presence of sodium
acetate under microwave irradiation to furnish new 1-(1H-benzo[d]
4.2. Pharmacological assay (in vitro anticancer activity)
4.2.1. Primary at a single dose (10 mM) full NCI 60 cell panel
All the selected compounds (16 in no.) were submitted to
imidazol-2-yl)-3-(5-(methyl
substituted)-1,3,4-oxadiazol-2-yl)
National Cancer Institute (NCI), USA for evaluating their in vitro
anticancer activity at single dose (10 mM) against full NCI 60 cell lines
propan-1-ones (IbeA7aee) (Fig. 2). Spectral data FT-IR, 1H NMR, 13
C
NMR and massspectra of the synthesized compounds wererecorded
and found to be in full agreement with the proposed structures. The
elemental analysis results were found to be within ꢀ0.4% of the
theoretical values. The substituent (R), irradiation time, NCS code
no., yield, melting point, molecular formula and molecular weight of
the newly synthesized compounds have been provided in Table 1.
panels representing full nine human systems as leukemia, mela-
noma and cancers of lung, colon, brain, breast, ovary, kidney and
prostate. The compounds added at a concentration (10 mM) and the
culture incubated for 48 h. End point determinations made with
a protein binding dye, sulforhodamine B. Results for each compound
were reported as a mean graph of the percent growth of the treated
cells and obtained result mentioned in Table 2.There after obtaining
the results for one dose assay, analysis of historical Developmental
Therapeutics Program (DTP) was performed and compound IbeA18
(NCS: 759205) satisfied pre-determined threshold inhibition
criteria and selected for NCI full panel five dose assay.
3. Pharmacological evaluation
3.1. In vitro anticancer assay [1,28e31]
The selected compounds among the newly synthesized
compounds were submitted for in vitro anticancer assay at National
Cancer Institute (NCI), USA against full NCI 60 cell lines panel
representing full nine human systems as leukemia, melanoma and
cancers of lung, colon, brain, breast, ovary, kidney and prostate.
Initially, structures of the synthesized compounds are submitted to
the NCI. The structures are generally selected for screening based
on their ability to add diversity to the NCI small molecule
compound collection. The structures with novel heterocyclic ring
systems are particularly given attention. The screening is a two-
stage process, beginning with the evaluation of all compounds
4.2.2. In vitro 5 dose full NCI 60 cell panel assay
In the second step, the selected compounds are evaluated
against all the 60 cell lines, representing nine tumor subpanels, at
10-fold dilutions of five different concentrations (0.01, 0.1, 1, 10 and
100 mM). The result are given in three calculated response param-
eters (GI50, TGI and LC50) for each cell line from log concentration vs
% growth inhibition curves. The GI50 value (growth inhibitory
activity) corresponds to the concentration of the compound causing
50% decrease in net cell growth, the TGI value (cytostatic activity) is
the concentration of the compound resulting in total growth inhi-
bition and LC50 value (cytotoxic activity) is the concentration of the
compound causing net 50% loss of initial cells at the end of the
incubation period of 48 h.
against the 60 cell lines at a single dose of 10 mM. The output from
the single dose screen was reported as a mean graph and available
for analysis by the COMPARE program. The compounds which

M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
857
Fig. 1. Protocol for synthesis of title compounds 1beA7e26
.
The compound IbeA18 (NCS: 759205) exhibited remarkable
anticancer activity against most of the tested cell lines with GI50
and 17.4
sensitivity profile toward Melanoma subpanel (GI50 value ranging
from 2.00 to 13.7 M), least for MDA-MB-435 (GI50, 2.00 M) and
maximum for UACC-257 (GI50, 13.7 M) cell line. All the tested
Leukemia cancer cell lines were found to be sensitive with not more
than 4.86 M concentrations of the tested compound. The highest
growth inhibitory activity was observed against the breast cancer
MDA-MB-468 cancer cell line with GI50 less than 1 M (0.797). The
mM, respectively. Obtained data revealed an obvious
values between 0.79 and 17.8
mM (Table 3). With regard to the
m
m
sensitivity against some individual cell lines, the compound showed
high activity against MDA-MB-468 (Breast Cancer), A498 (Renal
Cancer) and HOP-92 (Non-Small Cell Lung Cancer) with GI50 0.797,
1.53 and 1.77 and less sensitivity against SNB-75 (CNS Cancer), DU-
145 (Prostate Cancer) and SNB-19 (CNS Cancer) with GI50 18.7, 17.8
m
m
m

858
M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
a liquid paraffin bath in open capillary tubes and are uncorrected.
Progress of the reactions was monitored by using TLC plates (silica
gel G), Toluene: Ethyl acetate: Formic acid (5:4:1, v/v/v) and
benzene: acetone (9:1, v/v) used as solvent systems. The spots were
located by exposure to iodine vapors or under UV-light. Microwave
irradiation of reactions was done in
a scientific microwave
synthesizer (model No. CATA-R, Catalyst systems, India). 1H NMR
and 13C NMR spectra of the synthetic compounds were recorded on
Bruker spectropsin DPX-300 MHz in DMSO-d6/CDCl3; chemical
shift (d) values reported in parts per million (ppm) using tetrame-
thylsilane as internal reference. The splitting pattern abbreviations
are as follows: s, singlet; bs, broad singlet d, doublet; dd, double
doublet; t, triplet; m, multiplet. Mass spectra were recorded on
LCMS/MS (PerkineElmer and LABINDIA, Applied Biosystem) model
no. API 3000, presented as m/z. IR spectra were recorded on FT/IR
(Jasco), model no.410 using KBr pellets of the compounds.
Elemental analyses were performed on a PerkineElmer 240
analyzer and found to be in the range of ꢀ0.4% for each element
analyzed (C, H, N).
5.1.1. 4-(1H-Benzo[d]imidazol-2-yl)-4-oxobutanoic acid (1)
A solution of o-phenylene diamine (0.01 mol),
a-ketoglutaric
acid (equimolar; 0.01 mol) in methanol/water mixture (1:1) and
HCl (4N; 5 mL) was refluxed for 6 h and then left to cool to room
temperature. The NaOH solution (10% w/v) was added slowly to
neutralize the reaction mixture, when a solid mass precipitated out,
which was filtered, washed with water and recrystallized with
ethanol. Yield: 87%, m.p. 261e262 ꢁC, Rf ¼ 0.71 [toluene: ethyl
acetate: formic acid (5:4:1)]. IR (KBr, cmꢂ1): 3394 (OeH), 3326
(NeH), 3114 (CeH, AreH), 2972(CeH, CH2), 1728 (C]O), 1600 (C]
N), 1562 (C]C). 1H NMR (DMSO-d6): 12.97 (s, 1H, OH, D2O
exchangeable), 12.34 (s, 1H, NH, D2O exchangeable), 7.66 (d, 1H,
J ¼ 7.5 Hz, H-4, benzimidazole), 7.48 (t, 1H, J ¼ 7.2 Hz, H-7, benz-
imidazole), 7.27 (t, 2H, J ¼ 8.7 Hz, H-5,6 benzimidazole), 3.05 (t, 2H,
J ¼ 6.9 Hz, CH2), 2.76 (t, 2H, J ¼ 6.9 Hz, CH2). 13C NMR (DMSO-d6):
175.57 (C]O), 168.21 (C]O, COOH), 159.67 (C]N), 138.27, 132.21,
130.19, 128.57, 124.65, 123.64 (Ar-C), 34.23 (CH2, CH2CO), 31.36
(CH2, CH2COOH). Anal. calcd. for C11H10N2O3: C, 60.55; H, 4.62; N,
12.84. Found: C, 60.63; H, 4.65; N, 12.98. ESI-MS (m/z): 218 (Mþ).
Fig. 2. Protocol for synthesis of title compounds 1beA7aee
.
all remaining subpanel cell line showed maximum sensitive toward
tested compound with not more than 16.4 M concentrations. The
m
detailed results are mentioned in Table 3. The criterion for selec-
tivity of a compound depends upon the ratio obtained by dividing
the full panel MIDa (the average sensitivity of all cell lines toward
the test agent) by their individual subpanel MIDb (the average
sensitivity of all cell lines of a particular subpanel toward the test
agent). Ratios between 3 and 6 refer to moderate selectivity; ratio
greater than 6 indicates high selectivity toward the corresponding
cell line, while compounds not meeting either of these criteria rated
as non-selective. As per this criterion, compound in the study found
to be mild selective toward Leukemia cancer subpanel.
An analysis of results (structure activity relationship) indicated
that that electron donating groups like (eNH2, 2-chloro and 2,4-
dichloro) on the phenyl ring at position 5 of oxadiazole moiety
have high influence on anticancer activity and groups like
eN(C2H5)2, eOeC6H5 on methyl at position 5 of the oxadiazole ring
also resulted in compounds with improved anticancer activity.
Electron withdrawing groups like eCOOH and eC]O did not show
remarkable anticancer activity. Further studies to acquire more
information about quantitative structure activity relationship
(QSAR) of the compounds are in progress in our laboratory.
5.1.2. Ethyl- 4-(1H-benzo[d]imidazol-2-yl)-4-oxobutanoate (1a)
A solution of compound (1) (0.01 mol) in absolute ethanol
(25 mL) was refluxed for 10 h in presence of conc. H2SO4, (0.1 mL).
After completion of the reaction, it was cooled to room tempera-
ture, diluted with cold water and then neutralized with NaHCO3 to
pH 7. A precipitate formed which was filtered and crystallized from
ethanol. Yield: 81%, m.p. 253e254 ꢁC, Rf ¼ 0.60, [toluene: ethyl
acetate: formic acid (5:4:1)]. IR (KBr, cmꢂ1): 3330 (NeH), 3047
(CeH, Ar-H), 2958(CeH, CH2), 1718 (C]O), 1640 (C]N), 1570 (C]
C). 1H NMR (DMSO-d6): 12.51 (bs, 1H, NH, D2O exchangeable), 7.73
(d, 1H, J ¼ 7.8 Hz, H-4, benzimidazole), 7.54 (t, 1H, J ¼ 7.5 Hz, H-7,
benzimidazole), 7.12 (t, 2H, J ¼ 7.5 Hz, H-5,6, benzimidazole), 3.82
(q, 2H, J ¼ 5.4 Hz, CH2), 3.03 (t, 2H, J ¼ 7.2 Hz, CH2), 2.71 (t, 2H,
J ¼ 7.2 Hz, CH2), 1.15 (t, 3H, J ¼ 1.2 Hz, CH3). 13C NMR (DMSO-d6):
177.61(C]O), 165.37 (C]O, ester), 156.47(C]N), 138.32, 137.93,
129.13, 128.17, 124.55, 123.74 (AreC), 30.81 (CH2, CH2CO),
28.36(CH2, CH2COO), 69.07(CH2, ester), 20.31(CH3). Anal. calcd. for
C13H14N2O3: C, 63.40; H, 5.73; N, 11.38. Found: C, 62.77; H, 5.81; N,
11.35. ESI-MS (m/z): 246 (Mþ).
5. Experimental protocols
5.1.3. 4-(1H-Benzo[d]imidazol-2-yl)-4-oxobutane hydrazide (1b)
A solution of compound (Ia, 0.01 mol) and hydrazine hydrate
(0.015 mol) in ethanol (25 mL) was refluxed for 8 h. The reaction
mixture was cooled, a solid precipitate separated out, which was
filtered, dried and recrystallized from methanol. Yield: 85%, m.p.
5.1. Chemistry
All the reagents and solvents were obtained from S.D. Fine
chemicals or E. Merck Ltd. Melting points were determined on

M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
859
Table 1
Substituted value (eR), NCS code, yield, Rf-value, melting point, molecular formula/mol. weight and eluent mixture ratio of title compounds (1beA7e26) and (1beA7aee
)
.
Compd. No.
1beA7
eR
NCS codea
755135
ns
Yield
85
Rfꢂ value
0.65
Melting point (ꢁC)
223-224
Mol. Formula/Mol. weight
C13H11ClN4O2/290
Eluent mixture ratiob
9:1
8:2
1beA8
72
0.61
212-213
C16H17N5O2/311
1beA9
ns
82
0.73
233-235
C17H21N5O2/327
8:2
1beA10
1beA11
755139
759200
85
90
0.56
0.58
203-204
199-201
C17H13N5O2/319
C18H15N5O2/333
8:2
8:2
1beA12
1beA13
759201
759202
87
82
0.56
0.58
240-242
233-234
C19H15ClN4O2/366
C19H16N4O3/348
9:1
8:2
1beA14
1beA15
1beA16
1beA17
1beA18
1beA19
1beA20
1beA21
759203
759204
ns
70
80
75
72
94
85
83
87
0.43
0.64
0.55
0.51
0.53
0.52
0.60
0.50
246-247
217-218
221-222
211-213
227-228
217-219
241-242
209-210
C25H20N4O2/408
C14H11N5O2/281
C20H16N4O4/376
C14H12N4O2/268
C18H12Cl2N4O2/387
C18H14N4O2S/350
C15H14N4O2/282
C18H14N4O3/334
7:3
9:1
9:1
8:2
9:1
8:2
7:3
ns
759205
ns
ns
ns
8:2
(continued on next page)

860
M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
Table 1 (continued )
Compd. No.
eR
NCS codea
Yield
Rfꢂ value
Melting point (ꢁC)
Mol. Formula/Mol. weight
Eluent mixture ratiob
1beA22
1beA23
1beA24
1beA25
1beA26
ns
92
78
82
92
84
0.62
0.75
0.61
0.67
0.63
218-219
224-225
213-214
231-232
219-220
C18H15N5O3/349
C14H12N4O4/300
C13H12N4O2S/288
C19H16N4O3/348
C18H14N4O3/334
8:2
9:1
8:2
8:2
8:2
759206
ns
759207
759208
1beA7a
759209
78
0.49
242-243
C17H21N5O2/327
7:3
1beA7b
1beA7c
759210
759211
80
78
0.45
0.59
243-244
225-226
C25H21N5O2/423
C18H22N6O2/354
7:3
7:3
1beA7d
760445
759212
72
86
0.68
0.66
240-241
237-238
C21H15N5O4/401
C17H14N6O4/366
8:2
8:2
1beA7e
a
Some compounds were not selected (ns) for anticancer activity by the NCI.
Hexane and ethyl acetate mixture was used as eluent mixture.
b
241e242 ꢁC, Rf ¼ 0.53 [toluene: ethyl acetate: formic acid (5:4:1)].
IR (KBr, cmꢂ1): 3494 (NeH, NH2), 3359 (NeH, NH), 3151 (CeH,
AreH), 2947(CeH, CH2), 1691 (C]O), 1643 (C]N), 1600 (C]C).
1H NMR (DMSO-d6): 11.51 (s, 1H, NH, benzimidazole, D2O
exchangeable), 10.72 (s, 1H, NH, hydrazide, D2O exchangeable), 9.05
(bs, 2H, NH2, D2O exchangeable), 7.68 (d, 1H, J ¼ 7.8 Hz, H-4
benzimidazole), 7.50(t, 1H, J ¼ 7.2 Hz, H-7, benzimidazole), 7.27 (t,
2H, J ¼ 7.5 Hz, H-5,6 benzimidazole), 3.02 (t, 2H, J ¼ 7.5 Hz, CH2),
2.52 (t, 2H, J ¼ 7.8 Hz, CH2CO).13C NMR (DMSO-d6): 179.15 (C]O),
162.72 (C]O, hydrazide), 156.32 (C]N), 133.99, 132.28, 129.59,
128.45, 122.92, 116.85 (Ar-C), 30.17 (CH2, CH2CO), 22.27 (CH2,
hydrazide). Anal. calcd. for C11H12N4O2: C, 56.89; H, 5.21; N, 24.12.
Found: C, 56.73; H, 5.27; N, 24.15. ESI-MS (m/z): 232 (Mþ).
poured slowly onto crushed ice, neutralized with sodium bicar-
bonate solution. A solid mass precipitated out, which was filtered
and washed with excess quantity of water to remove the inorganic
component. The product was purified by flash chromatography
(hexane/ethyl acetate gradient, 8:2, concentration of gradient
varied compound to compound) to get pure compound (1beA7e26).
5.1.4.1. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(chloromethyl)-1,3,4-
oxadiazol-2-yl)propan-1-one (1beA7). IR (KBr, cmꢂ1): 3350 (NeH,
NH), 3020 (CeH, AreH), 2935(CeH, CH2), 1710 (C]O), 1682 (C]
N), 1574 (C]C), 1304 (NeN]C), 1164 (CeOeC asymmetric), 1028
(CeOeC, symmetric), 714 (CeCl). 1H NMR (CDCl3): 12.51 (s, 1H,
NH, D2O exchangeable), 7.80 (d, 1H,
J
¼
7.8 Hz, H-4,
benzimidazole), 7.48 (t, 1H, J ¼ 7.2 Hz, H-7, benzimidazole), 7.33(t,
2H, J ¼ 5.4 Hz, H-5,6, benzimidazole), 3.29 (t, 2H, J ¼ 6.9 Hz, CH2),
2.90 (t, 2H, J ¼ 6.6 Hz, CH2), 2.66 (s, 3H, CH3). 13C NMR (CDCl3):
169.45 (C]O), 157.21, 155.36 (2C, oxadiazole), 153.67 (C]N),
135.15, 133.21, 131.13, 127.53, 125.10, 125.84 (AreC), 50.01 (CH2,
CH2Cl), 38.75 (CH2, CH2CO), 27.41 (CH2). Anal. calcd./Found: [C,
53.71/53.62; H, 3.81/3.83; N, 19.27/19.41]. ESI-MS (m/z): 290 (Mþ).
5.1.4. General Procedure for synthesis of 1-(1H-Benzo[d]imidazol-2-
yl)-3-(1,3,4-oxadiazol-5 -substituted-2-yl) propan-1-one (1beA7e26
)
A
mixture of 4-(1H-benzo[d]imidazol-2-yl)-4-
oxobutanehydrazide (1b) (0.001 mol), an aliphatic/aromatic acid
(equimolar, 0.001 mol) and POCl3 (5 mL) was placed into a micro-
wave reaction vessel equipped with a magnetic stir bar. The reaction
vessel was placed into the scientific microwave synthesizer and
irradiated at a power level of 6 (60%, 420 W) for 6e13 min. After
completion of the reaction, checked by single-spot TLC using the
solvent systems; toluene: ethyl acetate: formic acid (5:4:1) or
benzene: acetone (9:1), the reaction mixture was cooled and
5.1.4.2. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(pyrrolidin-2-yl)-1,3,4-
oxadiazol-2-yl)propan-1-one (1beA8). IR (KBr, cmꢂ1): 3365 (NeH,
NH), 3030 (CeH, AreH), 2960 (CeH, CH2), 2824 (CeH, cyclic
CH2), 1690 (C]O), 1615 (C]N), 1542 (C]C), 1305 (NeN]C), 1130

M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
861
pyrrolidine). 13C NMR (CDCl3): 172.61 (C]O), 161.01, 158.63 (2C
oxadiazole), 155.21 (C]N), 132.17, 131.83, 131.03, 127.03, 124.71,
123.61 (AreC), 55.07, 53.17, 50.13, 44.51 (4C pyrrolidine), 38.07
(CH2, CH2CO), 30.41 (CH2). Anal. calcd./Found: [C, 61.72/61.83; H,
5.50/5.54; N, 22.49/22.37]. ESI-MS (m/z): 311 (Mþ).
Table 2
In vitro anticancer testing results of compound IbeA18 (NCS: 759205).
Panel
Cell line
Developmental therapeutics program
one dose mean graph value (10 M)
m
Growth percent
Growth inhibition
percent
5.1.4.3. 3-(5-(1-Amino-3-methylbutyl)-1,3,4-oxadiazol-2-yl)-1-(1H-
benzo[d]imidazol-2-yl) propan-1-one (1beA9). IR (KBr, cmꢂ1): 3484
(NeH, NH2), 3324 (NeH, NH), 3016 (CeH, AreH), 2952 (CeH, CH2),
1712 (C]O), 1635 (C]N), 1562 (C]C), 1328 (NeN]C), 1176
(CeOeC, asymmetric), 1020 (CeOeC, symmetric). 1H NMR
(CDCl3): 12.71 (s, 1H, NH, D2O exchangeable), 8.37 (s, 2H, NH2,
D2O exchangeable), 7.60 (d, 1H, J ¼ 7.5 Hz, H-4, benzimidazole),
7.41 (t, 1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.28 (t, 2H, J ¼ 7.5 Hz,
H-5,6, benzimidazole), 4.16 (t, 1H, J ¼ 7.2, CeH), 3.31 (t, 2H,
J ¼ 6.9 Hz, CH2), 2.87 (t, 2H, J ¼ 6.9 Hz, CH2), 1.49 (dd, 2H,
J ¼ 5.4 Hz, J ¼ 5.7 Hz, CH2), 1.36 (q, 1H, J ¼ 5.4 Hz, CH), 1.05 (d,
6H, J ¼ 1.2 Hz, 2xCH3). 13C NMR (CDCl3): 177.41 (C]O), 162.13,
161.51 (2C oxadiazole), 154.97 (C]N), 131.73, 131.01, 128.31,
127.07, 125.41, 124.37 (AreC), 56.07 (CH, CHeNH2), 49.17 (CH2,
CH2CH), 36.73 (CH2, CH2CO), 31.19 (CH, CHCH3), 27.41 (CH2),
25.61 (CH3, CHCH3). Anal. calcd./Found: [C, 62.37/61.79; H, 6.47/
6.41; N, 21.39/21.36]. ESI-MS (m/z): 327(Mþ).
Leukemia
CCRF-CEM
HL-60(TB)
K-562
MOLT-4
RPMI-8226
A549/ATCC
EKVX
HOP-62
HOP-92
NCI-H226
NCI-H23
NCI-H322M
NCI-H460
NCI-H522
COLO 205
HCC-2998
HCT-116
HCT-15
HT29
KM12
SW-620
SF-268
SF-295
SF-539
SNB-19
SNB-75
U251
47.29
ꢂ33.81a
2.22
20.71
30.87
69.67
60.96
71.65
6.67
83.71
61.94
75.97
77.33
49.76
62.65
65.88
22.07
81.50
9.39
28.69
35.81
68.09
59.47
95.24
75.09
63.36
52.50
55.09
79.33
55.79
ꢂ0.53a
59.44
82.67
66.98
78.62
42.34
57.56
66.38
58.75
81.15
57.16
63.29
64.83
73.82
58.42
81.30
59.83
81.55
61.18
63.99
52.05
53.14
82.16
44.72
38.32
52.71
133.81
97.78
79.29
69.13
30.33
39.04
28.35
93.33
16.29
38.06
24.03
22.67
50.24
37.35
34.12
77.93
18.50
90.61
71.31
64.19
31.91
40.53
4.76
24.91
36.64
47.50
44.91
20.67
44.21
100.53
40.56
17.33
33.02
21.38
57.66
42.44
33.62
41.25
18.85
42.84
36.71
35.17
26.18
41.58
18.70
40.17
18.45
38.82
36.01
47.95
46.86
17.84
55.28
61.68
Non-small cell
lung cancer
Colon cancer
CNS cancer
Melanoma
5.1.4.4. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(pyridin-2-yl)-1,3,4-
oxadiazol-2-yl)propan-1-one (1beA10). IR (KBr, cmꢂ1): 3374 (NeH,
NH), 3042 (CeH, AreH), 2934(CeH, CH2), 1720 (C]O), 1625 (C]N),
1560 (C]C), 1322 (NeN]C), 1152 (CeOeC, asymmetric), 988
(CeOeC, symmetric). 1H NMR (CDCl3): 12.57 (s, 1H, NH, D2O
exchangeable), 8.24e7.82 (m, 4H, pyridine), 7.70 (d, 1H, J ¼ 7.5 Hz,
H-4, benzimidazole), 7.45 (t, 1H, J ¼ 7.8 Hz, H-7, benzimidazole),
7.21 (t, 2H, J ¼ 7.5 Hz, H-5,6, benzimidazole), 3.17 (t, 2H, J ¼ 7.2 Hz,
CH2), 2.69 (t, 2H, J ¼ 7.2 Hz, CH2). 13C NMR (CDCl3): 175.63 (C]O),
165.37, 163.71 (2C oxadiazole), 153.43 (C]N), 138.23, 138.03, 134.17,
133.71, 131.83 (5C pyridine), 130.97, 130.07, 128.50, 127.61, 125.47,
124.93 (AreC), 33.43(CH2, CH2CO), 26.31(CH2). Anal. calcd./Found:
[C, 63.94/63.81; H, 4.10/4.17; N, 21.93/21.95]. ESI-MS (m/z): 319 (Mþ).
LOX IMVI
MALME-3M
M14
MDA-MB-435
SK-MEL-2
SK-MEL-28
SK-MEL-5
UACC-257
UACC-62
IGROV1
OVCAR-3
OVCAR-4
OVCAR-5
OVCAR-8
NCI/ADR-RES
SK-OV-3
786-0
A498
ACHN
CAKI-1
RXF 393
SN12C
TK-10
UO-31
PC-3
DU-145
MCF7
Ovarian cancer
Renal cancer
5.1.4.5. 3-(5-(2-Aminophenyl)-1,3,4-oxadiazol-2-yl)-1-(1H-benzo[d]
imidazol-2-yl)propan-1one (1beA11). IR (KBr, cmꢂ1): 3550 (NeH,
NH2), 3340 (NeH, NH), 3110 (CeH, AreH), 2970 (CeH, CH2), 1710
(C]O), 1660 (C]N), 1564 (C]C), 1314 (NeN]C), 1132 (CeOeC,
asymmetric), 1038 (CeOeC, symmetric). 1H NMR (DMSO-d6):
12.37 (s, 1H, NH, D2O exchangeable), 8.21 (s, 2H, NH2), 7.74 (d, 1H,
J ¼ 8.1.Hz, H-4, benzimidazole), 7.54 (t, 1H, J ¼ 7.5 Hz, H-7,
benzimidazole), 7.41 (t, 2H, J ¼ 7.5 Hz, H-5,6, benzimidazole),
7.32e7.02 (m, 4H, phenyl), 2.91 (t, 2H, J ¼ 7.2 Hz, CH2), 2.37 (t,
2H, J ¼ 6.9 Hz, CH2). 13C NMR (DMSO-d6): 179.13 (C]O), 164.83,
161.21 (2C oxadiazole), 155.71 (C]N), 138.01, 132.61, 131.13,
130.97, 130.11, 129.51, 128.57, 127.63, 122.47, 121.93, 119.27 (AreC),
40.75 (CH2, CH2CO), 25.81(CH2). Anal. calcd./Found: [C, 64.86/
64.78; H, 4.54/4.52; N, 21.01/21.13]. ESI-MS (m/z): 333(Mþ).
Prostate cancer
Breast cancer
MDA-MB-231
/ATCC
HS 578T
BT-549
T-47D
MDA-MB-468
Mean
82.33
56.35
51.96
ꢂ2.19a
55.13
17.67
43.65
48.04
5.1.4.6. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(2-chlorobenzyl)-1,3,4-
oxadiazol-2-yl)propan-1-one (1beA12). IR (KBr, cmꢂ1): 3326 (NeH),
3050 (CeH, AreH), 2890 (CeH, CH2),1724 (C]O),1630 (C]N),1506
(C]C), 1306 (NeN]C), 1150 (CeOeC, asymmetric), 1074 (CeOeC,
symmetric), 720 (CeCl). 1H NMR (DMSO-d6): 12.58 (s, 1H, NH, D2O
exchangeable), 7.81 (d, 1H, J ¼ 8.7 Hz, H-4, benzimidazole), 7.51 (t,
1H, J ¼ 7.2 Hz, H-7, benzimidazole), 7.48 (t, 2H, J ¼ 7.5 Hz, H-5,6,
benzimidazole), 7.29e7.09 (m, 4H, phenyl), 3.71 (bs, 2H, CH2),
3.01(t, 2H, J ¼ 6.9 Hz, CH2), 2.71 (t, 2H, J ¼ 6.9 Hz, CH2). 13C NMR
(DMSO-d6): 177.56 (C]O), 162.72, 159.86 (2C oxadiazole), 156.39
(C]N), 136.28, 135.59, 133.92, 129.45, 128.41, 125.74, 124.99,
123.63, 122.13, 121.65, 120.91, 118.76 (AreC), 35.83(CH2, CH2CO),
102.19
a
Negative (ꢂ) indicates the cell kill.
(CeOeC, asymmetric), 1018 (CeOeC, symmetric). 1H NMR (CDCl3):
12.77 (s, 1H, NH, benzimidazole, D2O exchangeable), 7.78 (d, 1H,
J ¼ 7.5 Hz, H-4, benzimidazole), 7.51 (t, 1H, J ¼ 7.2 Hz, H-7,
benzimidazole), 7.30 (t, 2H, J ¼ 8.7 Hz, H-5,6, benzimidazole),
3.92 (s, 1H, NH pyrrolidine, D2O exchangeable), 3.01 (t, 2H,
J ¼ 6.9 Hz, CH2), 2.74 (t, 2H, J ¼ 6.9 Hz, CH2), 2.30e1.19 (m, 7H,

862
M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
Table 3
Calculated values of GI50, TGI and LC50 of the cell lines, NCI: full cell lines, of compound IbeA18 (NCS: 759205).
Panel
Cell line
GI50
TGI (10ꢂ5M)
LC50 (10ꢂ4M)
Concentration per cell line (10ꢂ6M)
Subpanel MIDb
3.48
Leukemia
CCRF-CEM
HL-60(TB)
MOLT-4
RPMI-8226
SR
4.86
2.23
4.07
3.61
2.63
10.4
5.48
7.67
1.77
6.66
3.23
4.99
2.66
6.52
3.33
5.13
3.77
3.16
3.46
7.30
5.98
10.5
17.4
18.7
10.9
4.03
13.0
5.86
2.00
12.7
2.88
13.7
3.34
5.26
5.63
5.93
7.82
9.26
5.47
4.60
10.7
1.53
11.9
3.49
4.30
6.22
16.4
6.71
3.71
17.8
4.57
2.68
9.71
7.54
3.34
0.797
6.59
6.60
1.93
2.58
2.33
1.52
10.0
10.0
4.13
1.58
3.12
6.32
2.86
0.708
3.23
1.35
4.86
5.51
1.10
1.63
4.66
4.29
3.96
10.0
10.0
7.76
3.98
5.26
3.15
0.477
4.27
1.49
4.44
2.78
3.91
2.48
10.0
10.0
10.0
10.0
3.05
4.55
1.01
10.0
4.11
4.97
5.69
10.0
3.29
10.0
10.0
10.0
4.26
9.39
9.25
6.00
1.22
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
0.525
1.00
1.00
1.00
0.743
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
0.646
1.00
1.00
1.00
0.907
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
1.00
Non-small cell lung cancer
A549/ATCC
EKVX
5.74
4.00
HOP-62
HOP-92
NCI-H226
NCI-H23
NCI-H460
COLO 205
HCC-2998
HCT-116
HCT-15
HT29
KM12
SW-620
SF-268
SF-295
SF-539
SNB-19
SNB-75
U251
LOX IMVI
MALME-3M
M14
MDA-MB-435
SK-MEL-28
SK-MEL-5
UACC-257
UACC-62
IGROV1
OVCAR-3
OVCAR-4
OVCAR-5
OVCAR-8
NCI/ADR-RES
SK-OV-3
786-0
A498
ACHN
CAKI-1
RXF 393
SN12C
TK-10
UO-31
PC-3
DU-145
MCF7
Colon cancer
CNS cancer
Melanoma
11.79
7.18
Ovarian cancer
Renal cancer
6.28
7.65
Prostate cancer
Breast cancer
10.75
4.77
MDA-MB-231/ATCC
HS 578T
BT-549
T-47D
MDA-MB-468
MIDa
a
Mean graph midpoint (arithmetical mean value of treated cancer cell lines) representing the average sensitivity of all cell lines (full panel) toward the test agent in mM.
Average sensitivity of all cell lines of a particular subpanel toward the test agent in mM.
b
27.21 (CH2, oxadiazole), 24.86 (CH2). Anal. calcd./Found: [C, 62.21/
benzimidazole), 7.40e6.95 (m, 5H, phenyl), 3.76 (s, 2H, CH2), 3.10
(t, 2H, J ¼ 7.2 Hz, CH2), 2.67 (t, 2H, J ¼ 7.5 Hz, CH2). 13C NMR
(DMSO-d6): 171.53 (C]O), 164.02, 161.93 (2C oxadiazole), 158.15
(C]N), 147.31, 143.12, 140.32, 133.18, 130.15, 128.51, 126.23,
124.46, 123.72, 122.13, 121.89, 120.57 (AreC), 65.34 (CH2, CH2O),
36.92 (CH2, CH2CO), 25.57 (CH2). Anal. calcd./Found: [C, 65.51/
65.57; H, 4.63/4.51; N, 16.08/16.04]. ESI-MS (m/z): 348 (Mþ).
62.23; H, 4.12/4.17; N, 15.27/15.27]. ESI-MS (m/z): 366 (Mþ).
5.1.4.7. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(phenoxymethyl)-1,3,4-
oxadiazol-2-yl)propan-1-one (1beA13). IR (KBr, cmꢂ1): 3377 (NeH),
3029 (CeH, AreH), 2999(CeH, CH2), 1728 (C]O), 1600 (C]N), 1562
(C]C), 1384 (NeN]C), 1228 (CeOeC, asymmetric), 1078 (CeOeC,
symmetric). 1H NMR (DMSO-d6): 12.41 (s, 1H, NH, D2O
exchangeable), 7.80 (d, 1H, J ¼ 7.5.Hz, H-4 benzimidazole), 7.50 (t,
1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.47 (t, 2H, J ¼ 7.2 Hz, H-5,6,
5.1.4.8. 3-(5-Benzhydryl-1,3,4-oxadiazol-2-yl)-1-(1H-benzo[d]imida-
zol-2-yl)propan-1-one (1beA14). IR (KBr, cmꢂ1): 3328 (NeH), 3056

M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
863
(CeH, AreH), 2966(CeH, CH2), 1695 (C]O), 1608 (C]N), 1512 (C]
C), 1350 (NeN]C), 1203 (CeOeC, asymmetric), 1089 (CeOeC,
symmetric). 1H NMR (DMSO-d6): 12.45 (s, 1H, NH, D2O
exchangeable), 7.73 (d, 1H, J ¼ 7.8 Hz, H-4, benzimidazole), 7.50
(t, 1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.32 (t, 2H, J ¼ 7.2 Hz, H-
5,6 benzimidazole), 7.24e6.73 (m, 10H, phenyl), 4.42 (bs, H, CH),
3.08 (t, 2H, J ¼ 6.9 Hz, CH2), 2.73 (t, 2H, J ¼ 6.9 Hz, CH2). 13C NMR
(DMSO-d6): 175.15 (C]O), 162.72, 161.17 (2C oxadiazole), 156.45
(C]N), 144.73, 143.75, 142.53, 137.26, 134.13, 130.41, 129.21,
126.87, 125.42, 123.47, 122.37, 121.89, 121.13, 120.57, 118.46, 118.23,
116.76 (AreC), 49.27 (CH2, CH2, biphenyl), 40.12 (CH2, CH2O),
26.13 (CH2, CH2CO). Anal. calcd./Found: [C, 73.51/73.53; H, 4.94/
4.81; N, 13.72/13.78]. ESI-MS (m/z): 408 (Mþ).
benzimidazole), 7.63 (t, 1H, J ¼ 7.2 Hz, H-7, benzimidazole), 7.48
(t, 2H, J ¼ 7.5 Hz, H-5,6 benzimidazole), 7.37e7.01 (m, 3H,
phenyl), 3.17 (t, 2H, J ¼ 6.9 Hz, CH2), 2.66 (t, 2H, J ¼ 6.9 Hz, CH2).
13C NMR (DMSO-d6): 174.45 (C]O), 163.23, 160.13 (2C
oxadiazole), 154.35 (C]N), 138.24, 135.45, 133.21, 131.76, 128.51,
124.15, 123.63, 122.97, 122.05, 121.67, 120.53, 118.94 (AreC), 33.76
(CH2, CH2CO), 24.57 (CH2). Anal. calcd./Found: [C, 55.83/55.86; H,
3.12/3.37; N, 14.47/14.63]. ESI-MS (m/z): 387 (Mþ).
5.1.4.13. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(2-mercaptophenyl)-
1,3,4-oxadiazol-2-yl)propan-1-one (1beA19). IR (KBr, cmꢂ1): 3344
(NeH), 3028 (CeH, AreH), 2925 (CeH, CH2), 2570 (SeH), 1703
(C]O), 1613 (C]N), 1602 (C]C), 1336 (NeN]C), 1128 (CeOeC,
asymmetric), 1056 (CeOeC, symmetric). 1H NMR (DMSO-d6):
12.07 (s, 1H, NH, D2O exchangeable), 7.78 (d, 1H, J ¼ 7.8.Hz, H-4,
benzimidazole), 7.54 (t, 1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.32
(t, 2H, J ¼ 7.5 Hz, H-5,6, benzimidazole), 7.32e6.92 (m, 4H,
phenyl), 3.37 (s, 1H, SH), 2.89 (t, 2H, J ¼ 7.2 Hz, CH2), 2.61 (t, 2H,
J ¼ 6.9 Hz, CH2). 13C NMR (DMSO-d6): 169.85 (C]O), 160.89,
160.02 (2C oxadiazole), 153.74 (C]N), 134.53, 133.21, 130.66,
129.43, 127.74, 125.47, 124.97, 124.13, 122.91, 121.78, 120.63, 119.71
(AreC), 37.14 (CH2, CH2CO), 26.28 (CH2). Anal. calcd./Found: [C,
61.70/61.75; H, 4.03/4.16; N, 15.99/15.95]. ESI-MS (m/z): 350(Mþ).
5.1.4.9. 2-(5-(3-(1H-Benzo[d]imidazol-2-yl)-3-oxopropyl)-1,3,4-
oxadiazol-2-yl)acetonitrile (1beA15). IR (KBr, cmꢂ1): 3404 (NeH),
3029 (CeH, AreH), 2947 (CeH, CH2), 2270 (CN), 1715 (C]O),
1604 (C]C), 1594 (C]N), 1343 (NeN]C), 1256 (CeOeC,
asymmetric), 1083 (CeOeC, symmetric). 1H NMR (CDCl3): 11.91
(s, 1H, NH, D2O exchangeable), 7.78 (d, 1H, J ¼ 8.1 Hz, H-4,
benzimidazole), 7.49 (t, 1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.32
(t, 2H, J ¼ 3.9 Hz, H-5,6, benzimidazole), 1.82 (s, 2H, CH2), 3.31 (t,
2H, J ¼ 6.9 Hz, CH2), 2.92 (t, 2H, J ¼ 6.9 Hz, CH2). 13C NMR
(CDCl3): 175.57 (C]O), 162.72, 159.67 (2C oxadiazole), 154.96
(C]N), 138.27, 132.21, 130.19, 128.57, 124.65, 123.64 (AreC),
115.78 (CN), 34.23 (CH2, CH2O), 29.32 (CH2, CH2), 21.83 (CH2,
CH2CN). Anal. calcd./Found: [C, 59.78/59.76; H, 3.94/3.68; N,
24.90/24.97]. ESI-MS (m/z): 281(Mþ).
5.1.4.14. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(prop-1-enyl)-1,3,4-
oxadiazol-2-yl)propan-1-one (1beA20). IR (KBr,cmꢂ1): 3350 (NeH),
3070 (CeH, AreH), 3040 (CeH, aliphatic-H), 3010 (CeH, CH2]
CH2), 2899 (CeH, CH2), 1710 (C]O), 1657 (C]N), 1596 (C]C),
1290 (NeNeC), 1145 (CeOeC, asymmetric), 1014 (CeOeC,
symmetric). 1H NMR (CDCl3): 12.06 (s, 1H, NH, D2O
exchangeable), 7.73 (d, 1H, J ¼ 8.1.Hz, H-4, benzimidazole), 7.40 (t,
1H, J ¼ 6.6 Hz, H-7, benzimidazole), 7.27 (t, 2H, J ¼ 8.7 Hz, H-5,6,
benzimidazole), 4.18 (d, 1H, J ¼ 7.2 Hz, CH), 4.13 (m, 1H, CH), 1.28
(t, 3H, J ¼ 7.2 Hz, CH3), 3.23 (t, 2H, J ¼ 7.2 Hz, CH2), 2.85 (t, 2H,
J ¼ 6.6 Hz, CH2). 13C NMR (CDCl3): 170.37 (C]O), 163.91, 162.27
(2C oxadiazole), 156.15 (C]N), 133.14, 132.71, 129.23, 128.63,
127.56, 124.31, 123.97, 122.76 (AreC), 34.65 (CH2, CH2CO), 23.17
(CH2), 15.09 (CH3). Anal. calcd./Found: [C, 63.82/64.02; H, 5.00/
5.04; N, 19.85/19.88]. ESI-MS (m/z): 282 (Mþ).
5.1.4.10. 2-(5-(3-(1H-Benzo[d]imidazol-2-yl)-3-oxopropyl)-1,3,4-
oxadiazol-2-yl)phenyl acetate (1beA16). IR (KBr, cmꢂ1): 3317
(NeH), 3076 (CeH, AreH), 2970(CeH, CH2), 1718 (C]O), 1612
(C]C), 1556 (C]N), 1323 (NeN]C), 1264 (CeOeC, asymmetric),
1104 (CeOeC, symmetric). 1H NMR (CDCl3): 12.15 (s, 1H, NH, D2O
exchangeable), 7.81 (d, 1H, J ¼ 7.8.Hz, H-4, benzimidazole), 7.54
(t, 1H, J ¼ 7.8 Hz, H-7, benzimidazole), 7.36 (t, 2H, J ¼ 7.5 Hz, H-
5,6, benzimidazole), 7.30e6.68 (m, 4H, phenyl), 2.24 (s, 3H, CH3),
2.91 (t, 2H, J ¼ 6.9 Hz, CH2), 2.53 (t, 2H, J ¼ 6.9 Hz, CH2). 13C NMR
(CDCl3): 176.68 (C]O), 169.87 (C]O, COCH3), 162.97, 161.77 (2C
oxadiazole), 155.67 (C]N), 150.02, 146.34, 140.13, 138.26, 130.31,
128.25, 125.32, 124.56, 123.72, 123.13, 122.92, 121.43 (AreC), 41.25
(CH2, CH2CO), 24.15 (CH2), 17.78 (CH3). Anal. calcd./Found: [C,
63.82/64.96; H, 4.28/4.16; N, 14.89/14.87]. ESI-MS (m/z): 376 (Mþ).
5.1.4.15. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(2-hydroxyphenyl)-
1,3,4-oxadiazol-2-yl)propan-1-one (1beA21). IR (KBr, cmꢂ1): 3396
(OeH), 3310 (NeH), 3105 (CeH, AreH), 2891 (CeH, CH2), 1700
(C]O), 1640 (C]N), 1610 (C]C), 1297 (NeN]C), 1204 (CeOeC,
asymmetric), 1040 (CeOeC, symmetric). 1H NMR (DMSO-d6):
11.87 (s, 1H, NH, D2O exchangeable), 7.92 (d, 1H, J ¼ 8.1 Hz, H-4,
benzimidazole), 7.51 (t, 1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.35
(t, 2H, J ¼ 7.5 Hz, H-5,6, benzimidazole), 7.27-7.01 (m, 4H,
phenyl), 6.51 (s, 1H, OH), 3.01 (t, 2H, J ¼ 6.9 Hz, CH2), 2.73 (t, 2H,
J ¼ 7.2 Hz, CH2). 13C NMR (DMSO-d6): 176.23(C]O), 166.41,
162.92(2C oxadiazole), 157.31(C]N), 148.12, 130.15, 129.32,
128.74, 126.51, 124.67, 123.87, 123.05, 122.78, 121.57, 120.43 (Ar-C),
36.71(CH2, CH2CO), 24.15(CH2). Anal. calcd./Found: [C, 64.66/
64.63; H, 4.22/4.31; N, 16.76/16.79]. ESI-MS (m/z): 334 (Mþ).
5.1.4.11. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-vinyl-1,3,4-oxadiazol-2-
yl)propan-1-one (1beA17). IR (KBr, cmꢂ1): 3356 (NeH), 3089 (CeH,
AreH), 2983 (CeH, H2C]CH2), 2945 (CeH, CH2), 1709 (C]O), 1643
(C]C), 1503 (C]N), 1299 (NeN]C), 1205 (CeOeC, asymmetric),
1076 (CeOeC, symmetric). 1H NMR (CDCl3): 11.94 (s, 1H, NH, D2O
exchangeable), 7.81 (d, 1H, J ¼ 8.1 Hz, H-4, benzimidazole), 7.51
(t, 1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.30 (t, 2H, J ¼ 6.3 Hz, H-
5,6, benzimidazole), 4.21 (t, J ¼ 7.2 Hz, H, CH), 4.14 (d, J ¼ 7.2 Hz,
2H, CH2), 3.32(t, 2H, J ¼ 7.2 Hz, CH2), 2.91 (t, 2H, J ¼ 5.7 Hz, CH2).
13C NMR (CDCl3): 177.58 (C]O), 163.72, 159.83 (2C oxadiazole),
156.32 (C]N), 133.99, 132.28, 129.59, 128.45 (AreC), 122.92,
116.85 (2C, CH2]CH2), 33.17 (CH2, CH2CO), 22.27 (CH2). Anal.
calcd./Found: [C, 62.68/62.63; H, 4.51/5.03; N, 20.88/20.56]. ESI-
MS (m/z): 268 (Mþ).
5.1.4.16. 3-(5-(4-Amino-2-hydroxyphenyl)-1,3,4-oxadiazol-2-yl)-1-
(1H-benzo[d]imidazol-2-yl) propan-1-one (1beA22). IR (KBr, cmꢂ1):
3452 (NeH, NH2), 3390 (OeH), 3305 (NeH, NH), 3055 (CeH, AreH),
2931 (CeH, CH2), 1790 (C]O), 1654 (C]N), 1570 (C]C), 1317
(NeN]C), 1140 (CeOeC, asymmetric), 1015 (CeOeC, symmetric).
1H NMR (DMSO-d6): 12.09 (s, 1H, NH, D2O exchangeable), 7.78 (d,
1H, J ¼ 7.8 Hz, H-4, benzimidazole), 7.51 (t, 1H, J ¼ 7.5 Hz, H-7
benzimidazole), 7.30 (t, 2H, J ¼ 7.5 Hz, H-5,6, benzimidazole),
7.24e6.81 (m, 3H, phenyl), 6.71 (s, 2H, NH2), 6.03 (s, 1H, OH), 3.11
(t, 2H, J ¼ 7.2 Hz, CH2), 2.57 (t, 2H, J ¼ 7.2 Hz, CH2). 13C NMR
5.1.4.12. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(2,4-dichlorophenyl)-
1,3,4-oxadiazol-2-yl)propan-1-one (1beA18). IR (KBr, cmꢂ1): 3336
(NeH), 3093 (CeH, AreH), 2900 (CeH, CH2), 1685 (C]O), 1654
(C]N), 1627 (C]C), 1373 (NeN]C), 1161 (CeOeC, asymmetric),
1060 (CeOeC, symmetric), 763 (CeCl). 1H NMR (DMSO-d6): 11.99
(s, 1H, NH, D2O exchangeable), 7.87 (d, 1H, J ¼ 8.1.Hz, H-4,

864
M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
(DMSO-d6): 177.31 (C]O), 163.23, 162.12 (2C oxadiazole), 153.17
(C]N), 150.13, 147.32, 146.63, 130.14, 127.14, 126.53, 123.75,
122.78, 122.17, 121.53, 120.37, 119.63 (AreC), 38.85 (CH2, CH2CO),
27.53(CH2). Anal. calcd./Found: [C, 61.89/61.85; H, 4.33/4.41; N,
20.05/21.03]. ESI-MS (m/z): 349 (Mþ).
Found: [C, 64.66/64.71; H, 4.22/4.17; N, 16.76/16.89]. ESI-MS (m/
z): 334 (Mþ).
5.1.5. General procedure for synthesis of 1-(1H-benzo[d]imidazol-2-
yl)-3-(5-(methyl substituted)-1,3,4-oxadiazol-2-yl)propan-1-one
(1beA7aee
)
5.1.4.17. 2-(5-(3-(1H-Benzo[d]imidazol-2-yl)-3-oxopropyl)-1,3,4-
oxadiazol-2-yl)acetic acid (1beA23). IR (KBr, cmꢂ1): 3413 (OeH),
3301 (NeH), 3012 (CeH, AreH), 2850 (CeH, CH2), 1728 (C]O),
1670 (C]N), 1608 (C]C), 1384 (NeN]C), 1180 (CeOeC,
asymmetric), 1029 (CeOeC, symmetric). 1H NMR (DMSO-d6):
11.48 (s, 1H, NH, D2O exchangeable), 10.37 (s, 1H, OH), 7.78 (d, 1H,
J ¼ 8.1.Hz, H-4, benzimidazole), 7.48 (t, 1H, J ¼ 7.5 Hz, H-7,
benzimidazole), 7.26 (t, 2H, J ¼ 7.5 Hz, H-5,6, benzimidazole),
3.88 (bs, 2H, CH2), 3.08 (t, 2H, J ¼ 6.9 Hz, CH2), 2.68 (t, 2H,
J ¼ 6.9 Hz, CH2). 13C NMR (DMSO-d6): 178.67 (C]O), 169.43 (C]
O, COOH), 165.37, 163.28 (2C oxadiazole), 157.53 (C]N), 129.17,
128.45, 124.31, 123.74, 121.58, 120.78 (AreC), 41.13 (CH2, CH2CO),
36.23 (CH2, CH2COOH), 25.37 (CH2). Anal. calcd./Found: [C, 56.00/
56.03; H, 4.03/4.15; N, 18.66/18.67]. ESI-MS (m/z): 300 (Mþ).
The suspension of compound (1beA7) (0.003 mol) and
a secondary amine (equimolar; 0.003 mol) in absolute ethanol
(10 mL) was placed into a microwave reaction vessel equipped with
a magnetic stir bar, adding sodium acetate (0.001 mol) into
microwave reaction vessel equipped with a magnetic stir bar. The
reaction vessel was placed into the scientific microwave synthe-
sizer (model No. CATA-R, Catalyst systems, India) and irradiated at
a power level of 5 (50%, 350 W) for 8e13 min. The completion of
reaction was checked by single-spot TLC by using toluene: ethyl
acetate: formic acid (5:4:1) and benzene: acetone (9:1) solvent
system. The reaction mixture was cooled, poured onto crushed ice
and acidified with glacial acetic acid to yield a solid mass. It was
filtered and washed with water to remove the inorganic compo-
nents. The product was purified by flash chromatography (hexane/
ethyl acetate gradient, 8:2, concentration of gradient varied
compound to compound) to get pure compound (1beA7aee).
5.1.4.18. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(mercaptomethyl)-
1,3,4-oxadiazol-2-yl)propan-1-one (1beA24). IR (KBr, cmꢂ1): 3307
(NeH), 3076 (CeH, AreH), 2981 (CeH, CH2), 2490 (SeH), 1704
(C]O), 1650 (C]N), 1614 (C]C), 1376 (NeN]C), 1192 (CeOeC,
asymmetric), 1046 (CeOeC, symmetric). 1H NMR (CDCl3): 11.76
(s, 1H, NH, D2O exchangeable), 7.78 (d, 1H, J ¼ 8.7 Hz, H-4,
benzimidazole), 7.49 (t, 1H, J ¼ 7.8 Hz, H-7, benzimidazole), 7.33
(t, 2H, J ¼ 6.3 Hz, H-5,6, benzimidazole), 4.16 (d, J ¼ 7.2 Hz, 2H,
CH2), 3.34 (t, 2H, J ¼ 7.2 Hz, CH2), 2.92 (t, 2H, J ¼ 6.9 Hz, CH2),
1.27 (t, J ¼ 7.2 Hz, 1H, HS). 13C NMR (CDCl3): 173.42 (C]O),
160.97, 160.24 (2C oxadiazole), 155.33(C]N), 132.59, 131.92,
129.86, 128.50, 123.39, 116.00 (AreC), 51.80 (CH2, CH2SH),
29.80(CH2, CH2CO), 28.06 (CH2). Anal. calcd./Found: [C, 54.15/
54.16; H, 4.20/4.27; N, 19.43/19.31]. ESI-MS (m/z): 288 (Mþ).
5.1.5.1. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-((diethylamino)methyl)-
1,3,4-oxadiazol-2-yl)propan-1-one (1beA7a). IR (KBr, cmꢂ1): 3309
(NeH), 3072 (CeH, AreH), 2950 (CeH, CH2), 2873 (CeH, CH3),
1715 (C]O), 1670 (C]N), 1547 (C]C), 1373 (NeN]C), 1184
(CeOeC, asymmetric), 1077 (CeOeC, symmetric). 1H NMR
(CDCl3): 12.04 (s, 1H, NH, D2O exchangeable), 7.73 (d, 1H,
J ¼ 7.8.Hz, H-4, benzimidazole), 7.42 (d, 1H, J ¼ 11.7 Hz, H-7,
benzimidazole), 7.28 (t, 2H, J ¼ 8.7 Hz, J ¼ 9.6 Hz, H-5,6,
benzimidazole), 4.13 (q, 4H, J ¼ 5.4 Hz, 2xCH2), 3.23 (t, 2H,
J ¼ 6.9 Hz, CH2), 2.87 (t, 2H, J ¼ 6.6 Hz, CH2), 2.60 (s, 2H, CH2),
1.25 (t, 6H, J ¼ 1.2 Hz, 2xCH3). 13C NMR (CDCl3): 171.50 (C]O),
165.11, 161.19 (2C oxadiazole), 155.02 (C]N), 132.83, 131.96,
129.86, 128.52, 124.73, 123.48 (AreC), 60.24 (CH2, CH2N), 30.21
(CH2, CH2CO), 28.08 (CH2), 14.60 (CH3). Anal. calcd./Found: [C,
62.37/63.87; H, 6.47/6.51; N, 21.39/21.35]. ESI-MS (m/z): 327(Mþ).
5.1.4.19. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-(4-methoxyphenyl)-
1,3,4-oxadiazol-2-yl)propan-1-one (1beA25). IR (KBr, cmꢂ1): 3386
(NeH), 3082 (CeH, AreH), 2974 (CeH, CH2), 2758 (CeH, CH3),
1716 (C]O), 1624 (C]N), 1608 (C]C), 1384 (NeN]C), 1161
(CeOeC, asymmetric), 1022 (CeOeC, symmetric). 1H NMR
(DMSO-d6): 11.88 (s, 1H, NH, D2O exchangeable), 8.30e7.84 (m,
4H, phenyl), 7.70 (d, 1H, J ¼ 7.5.Hz, H-4, benzimidazole), 7.48 (t,
1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.27 (t, 2H, J ¼ 7.8 Hz, H-5,6,
benzimidazole), 4.15 (s, 3H, CH3), 3.21 (t, 2H, J ¼ 6.9 Hz, CH2),
2.83 (t, 2H, J ¼ 6.9 Hz, CH2). 13C NMR (DMSO-d6): 176.49 (C]O),
164.73, 161.48 (2C oxadiazole), 158.12 (C]N), 151.32, 133.52,
132.23, 130.63, 129.83, 128.74, 124.53, 123.39, 122.30, 121.43,
120.34 (AreC), 45.21 (CH3, OCH3), 32.67 (CH2, CH2CO), 25.47
(CH2). Anal. calcd./Found: [C, 65.51/C, 65.61; H, 4.63/4.75; N,
16.08/16.12]. ESI-MS (m/z): 348 (Mþ).
5.1.5.2. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-((diphenylamino)
methyl)-1,3,4-oxadiazol-2-yl) propan-1-one (1beA7b). IR (KBr,
cmꢂ1): 3290 (NeH), 3067 (CeH, AreH), 2957 (CeH, CH2), 1690
(C]O), 1654 (C]N), 1583 (C]C), 1287 (NeN]C), 1162 (CeOeC,
asymmetric), 1027 (CeOeC, symmetric). 1H NMR (DMSO-d6):
12.13 (s, 1H, NH, D2O exchangeable), 7.92 (d, 1H, J ¼ 8.7 Hz, H-4,
benzimidazole), 7.70 (d, 1H, J ¼ 7.8 Hz, H-7, benzimidazole), 7.48
(t, 2H, J ¼ 7.5 Hz, H-5,6, benzimidazole), 7.27e6.87 (m, 10H,
phenyl), 3.72 (s, 2H, CH2), 3.16 (t, 2H, J ¼ 7.2 Hz, CH2), 2.54 (t, 2H,
J ¼ 7.2 Hz, CH2). 13C NMR (DMSO-d6): 173.43 (C]O), 163.51,
162.73 (2C oxadiazole), 157.23 (C]N), 150.43, 149.27, 148.03,
133.17, 132.56, 128.12, 127.53, 127.02, 124.35, 123.43, 122.73,
120.47 (AreC), 57.48 (CH2, CH2N), 33.71 (CH2, CH2CO), 25.32
(CH2). Anal. calcd./Found: [C, 70.91/70.93; H, 5.00/5.07; N, 16.54/
16.68]. ESI-MS (m/z): 423(Mþ).
5.1.4.20. 1-(1H-benzo[d]imidazol-2-yl)-3-(5-(4-hydroxyphenyl)-
1,3,4-oxadiazol-2-yl)propan-1-one (1beA26). IR (KBr, cmꢂ1): 3420
(OeH), 3325 (NeH), 3043 (CeH, AreH), 2936 (CeH, CH2), 1697
(C]O), 1634 (C]N), 1585 (C]C), 1327 (NeN]C), 1176 (CeOeC,
asymmetric), 1058 (CeOeC, symmetric). 1H NMR (DMSO-d6):
12.05 (s, 1H, NH, D2O exchangeable), 8.11e7.85 (m, 4H, phenyl),
7.68 (d, 1H, J ¼ 7.5.Hz, H-4, benzimidazole), 7.51 (t, 1H, J ¼ 7.5 Hz,
H-7, benzimidazole), 7.27 (t, 2H, J ¼ 8.1 Hz, H-5,6, benzimidazole),
6.71 (s, 1H, OH), 3.09 (t, 2H, J ¼ 6.9 Hz, CH2), 2.75 (t, 2H,
J ¼ 6.9 Hz, CH2). 13C NMR (DMSO-d6): 171.93 (C]O), 162.87,
160.68 (2C oxadiazole), 155.27 (C]N), 150.16, 130.53, 129.53,
128.75, 125.34, 124.31, 123.91, 123.16, 122.70, 121.35, 120.53,
120.05 (AreC), 36.21 (CH2, CH2CO), 26.43 (CH2). Anal. calcd./
5.1.5.3. 1-(1H-Benzo[d]imidazol-2-yl)-3-(5-((4-methylpiperazin-1-
yl)methyl)-1,3,4-oxadiazol-2 yl) propan-1-one (1beA7c). IR (KBr,
cmꢂ1): 3317 (NeH), 3015 (CeH, AreH), 2941 (CeH, CH2), 2868
(CeH, CH3), 1711 (C]O), 1645 (C]N), 1576 (C]C), 1363 (NeN]
C), 1183 (CeOeC, asymmetric), 1057 (CeOeC, symmetric). 1H
NMR (DMSO-d6): 11.45 (s, 1H, NH, D2O exchangeable), 7.84 (d, 1H,
J ¼ 7.5 Hz, H-4, benzimidazole), 7.54 (d, 1H, J ¼ 7.5 Hz, H-7,
benzimidazole), 7.41 (t, 2H, J ¼ 7.8 Hz, H-5,6, benzimidazole), 3.74
(s, 2H, CH2), 3.59 (t, 4H, J ¼ 7.8 Hz, 2xCH2, piperazine), 3.15 (t, 4H,
J ¼ 6.9 Hz, 2xCH2-NCH3, piperazine), 3.01 (t, 2H, J ¼ 7.2 Hz, CH2),

M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
865
2.77 (t, 2H, J ¼ 7.2 Hz, CH2), 1.91 (s, 3H, CH3). 13C NMR (DMSO-d6):
175.31 (C]O), 165.32, 163.84 (2C oxadiazole), 156.13 (C]N), 131.52,
128.25, 124.57, 123.61, 123.04, 122.81 (AreC), 61.23 (CH2, CH2N),
54.27, 54.13, 51.83, 51.09 (4C piperazine), 43.71 (CH3), 35.02 (CH2,
CH2CO), 26.51 (CH2). Anal. calcd./Found: [C, 61.00/61.09; H, 6.26/
6.27; N, 23.71/23.83]. ESI-MS (m/z): 354 (Mþ).
dimethyl sulfoxide at 400-fold the desired final maximum test
concentration and stored frozen prior to use. At the time of drug
addition, an aliquot of frozen concentrate was thawed and diluted
to twice the desired final maximum test concentration with
complete medium containing 50
four, 10-fold or ½ log serial dilutions are made to provide a total of
five drug concentrations plus control. Aliquots of 100 L of these
different drug dilutions added to the appropriate microtiter wells
mg/mL gentamicin. Additional
m
5.1.5.4. 1-((5-(3-(1H-Benzo[d]imidazol-2-yl)-3-oxopropyl)-1,3,4-
oxadiazol-2-yl)methyl)indoline ꢂ2,3-dione (1beA7d). IR (KBr,
cmꢂ1): 3293 (NeH), 3041 (CeH, AreH), 2957 (CeH, CH2), 1716
(C]O), 1687 (C]N), 1572 (C]C), 1338 (NeN]C), 1168 (CeOeC,
asymmetric), 1052 (CeOeC, symmetric). 1H NMR (DMSO-d6):
11.95 (s, 1H, NH, D2O exchangeable), 8.74-7.52 (m, 4H, phenyl),
7.64 (d, 1H, J ¼ 7.5.Hz, H-4, benzimidazole), 7.51 (d, 1H, J ¼ 7.5 Hz,
H-7, benzimidazole), 7.30 (t, 2H, J ¼ 7.8 Hz, H-5,6, benzimidazole),
4.36 (s, 2H, CH2), 3.07 (t, 2H, J ¼ 7.2 Hz, CH2), 2.63 (t, 2H,
J ¼ 7.2 Hz, CH2). 13C NMR (DMSO-d6): 177.65 (C]O, COCH2),
175.23 (CO), 170.31 (C]O, CON), 161.83, 159.75 (2C oxadiazole),
154.97 (C]N), 147.23, 135.76, 133.63, 132.23, 124.35, 124.02,
123.43, 122.73, 122.16, 121.43, 120.73 (AreC), 56.89 (CH2, CH2N),
34.85 (CH2, CH2CO), 26.53 (CH2). Anal. calcd./Found: [C, 62.84/
62.87; H, 3.77/3.73; N, 17.45/17.52]. ESI-MS (m/z): 401(Mþ).
already containing 100
drug concentrations.
mL of medium, resulting in the required final
Following drug addition, the plates were incubated for an
additional 48 h at 37 ꢁC, 5% CO2, 95% air, and 100% relative humidity.
For adherent cells, the assay terminated by the addition of cold TCA.
Cells fixed in situ by the gentle addition of 50 mL of cold 50% (w/v)
TCA (final concentration, 10% TCA) and incubated for 60 min at 4 ꢁC.
The supernatant was discarded, and the plates washed five times
with tap water and then air dried. Sulforhodamine B (SRB) solution
(100 mL) at 0.4% (w/v) in 1% acetic acid was added to each well and
plates incubated for 10 min at room temperature. After staining,
unbound dye removed by washing five times with 1% acetic acid
and then the plates air dried. Bound stain subsequently solubilized
with 10 mM trizma base and the absorbance was read on an
automated plate reader at a wavelength of 515 nm. For suspension
cells, the methodology was same except that the assay terminated
by fixing settled cells at the bottom of the wells by gently adding
5.1.5.5. 3-((5-(3-(1H-Benzo[d]imidazol-2-yl)-3-oxopropyl)-1,3,4-
oxadiazol-2-yl)methyl) pyrimi-dine-2,4(1H,3H)-dione (1beA7e). IR
(KBr, cmꢂ1): 3345 (NeH), 3028 (CeH, AreH), 2911 (CeH, CH2), 1703
(C]O), 1681 (C]N), 1578 (C]C), 1316 (NeN]C), 1193 (CeOeC,
asymmetric), 1034 (CeOeC, symmetric). 1H NMR (DMSO-d6):
11.00 (s, 1H, NH, benzimidazole, D2O exchangeable), 10.81 (s, 1H,
50 mL of 80% TCA (final concentration, 16% TCA). Using the seven
absorbance measurements [time zero, (Tz), control growth, (C) and
test growth in the presence of drug at the five concentration levels
(Ti)], the percentage growth was calculated at each of the drug
concentrations levels using the following formula:
NH, D2O exchangeable), 7.81 (d, 1H,
J
¼
7.8 Hz, H-4,
benzimidazole), 7.54 (d, 1H, J ¼ 7.5 Hz, H-7, benzimidazole), 7.27
(t, 2H, J ¼ 7.5 Hz, H-5,6 benzimidazole), 4.32 (s, 2H, CH2), 3.23 (t,
2H, J ¼ 7.2 Hz, CH2), 2.57 (t, 2H, J ¼ 6.9 Hz, CH2). 13C NMR (DMSO-
d6): 173.51 (C]O), 170.38 (CO, CON), 159.02 (C]O, NCON), 164.75,
162.86 (2C, oxadiazole), 156.46 (C]N), 132.45, 131.64, 129.84,
124.76, 123.43, 122.53, 121.43, 118.83 (AreC), 58.96 (CH2, CH2N),
38.54 (CH2, CH2CO), 28.57 (CH2). Anal. calcd./Found: [C, 55.74/
55.71; H, 3.85/3.85; N, 22.94/22.82]. ESI-MS (m/z): 366 (Mþ).
½ðTi ꢂ TzÞ=ðC ꢂ TzÞꢃ
ꢄ 100 for concentrations for whichTi>= ¼ Tz
½ðTi ꢂ TzÞ=Tzꢃ ꢄ 100 for concentrations for which Ti < Tz:
Three dose response parameters (GI50, TGI and LC50) were
calculated for each experimental agent. Growth inhibition of 50%
(GI50) calculated from [(TieTz)/(CeTz)] ꢄ 100 ¼ 50, which was the
drug concentration resulting in a 50% reduction in the net protein
increase (as measured by SRB staining) in control cells during the
drug incubation. The drug concentration resulting in total growth
inhibition (TGI) calculated from Ti ¼ Tz. The LC50 (concentration of
drug resulting in a 50% reduction in the measured protein at the
end of the drug treatment as compared to that at the beginning)
indicating a net loss of cells following treatment was calculated
from [(TieTz)/Tz] ꢄ 100 ¼ ꢂ50. Values were calculated for each of
these three parameters, if the level of activity was reached;
however, if the effect is not reached or was exceeded, the value for
that parameter expressed as greater or less than the maximum or
minimum concentration tested.
5.2. In vitro anticancer activity [1,28e31]
The compounds were evaluated for their in vitro anticancer
activity at National Cancer Institute (NCI), USA against full NCI 60
cell lines panel representing on full nine human systems as
leukemia, melanoma and cancers of lung, colon, brain, breast,
ovary, kidney and prostate in accordance with their applied
protocol (used SRB assay). Sixteen compounds among the twenty-
five newly synthesized compounds were selected for anticancer
evaluation. The screening is a two-stage process, beginning with
the evaluation of all compounds against the 60 cell lines at a single
dose of 10 mM. The output from the single dose screen was reported
as a mean graph and available for analysis by the COMPARE
program. Compounds that exhibited significant growth inhibition
were evaluated against the 60 cell panel at five concentration
levels. The human tumor cell lines of the cancer screening panel
were grown in RPMI 1640 medium containing 5% fetal bovine
serum and 2 m ML-glutamine. For a typical screening experiment,
6. Conclusion
Twenty-five new benzimidazoles bearing 1,3,4-oxadiazole
nucleus were successfully synthesized under microwave irradiation
in good yields. The compounds were evaluated for their in vitro
anticancer activity against full NCI 60 cell lines panel. The screening
result data obtained from NCI indicated that the compounds 1beA7,
1beA11, 1beA12, 1beA13, 1beA14, 1beA15, 1beA23, 1beA7a, 1beA7b
and 1beA7c exhibited good percentage growth inhibition of human
cancer cell lines, and compound 1beA18 (NSC: 759205),1-(1H-benzo
[d] imidazol-2-yl)-3-(5-(2,4-dichlorophenyl)-1,3,4-oxadiazol-2-yl)
propan-1-one, showed maximum growth inhibition and found to be
the most active candidate of the series (lead compound). On the basis
cells inoculated into 96 well microtiter plates in 100 mL at plating
densities ranging from 5000 to 40,000 cells/well depending on the
doubling time of individual cell lines. After cell inoculation, the
microtiter plates incubated at 37 ꢁC, 5% CO2, 95% air and 100%
relative humidity for 24 h prior to addition of experimental drugs.
After 24 h, two plates of each cell line fixed in situ with TCA, to
represent a measurement of the cell population for each cell line at
the time of drug addition (Tz). Experimental drugs solubilized in

866
M. Rashid et al. / European Journal of Medicinal Chemistry 54 (2012) 855e866
[6] M.M. Ramla, M.A. Omar, A.M.M. El-Khamry, H.I.E. Diwani, Bioorg. Med. Chem.
14 (2006) 7324e7332.
[7] M.M. Ramla, M.A. Omar, H. Tokuda, H.I.E. Diwani, Bioorg. Med. Chem. 15
(2007) 6489e6496.
[8] Y. Li, C. Tan, C. Gao, C. Zhang, X. Luan, X. Chen, H. Liu, Y. Chen, Y. Jiang, Bioorg.
Med. Chem. 19 (2011) 4529e4535.
[9] H.M. Refaat, Eur. J. Med. Chem. 45 (2010) 2949e2956.
[10] S. Demirayak, I. Kayagil, L. Yurttas, Eur. J. Med. Chem. 46 (2011) 411e416.
[11] E. Moriarty, M. Carr, S. Bonham, M.P. Carty, F. Aldabbagh, Eur. J. Med. Chem. 45
(2010) 3762e3769.
[12] S.M. Sondhi, R. Rani, J. Singh, P. Roy, S.K. Agrawal, A.K. Saxena, Bioorg. Med.
Chem. Lett. 20 (2010) 2306e2310.
[13] S. Demirayak, A. Usama, A.C. Mohsen, K. Agri, Eur. J. Med. Chem. 37 (2002)
255e260.
[14] D. Sharma, B. Narasimhan, P. Kumar, V. Judge, R. Narang, E.D. Clercq,
J. Balzarini, J. Enzy. Inh. Med. Chem. 24 (2009) 1161e1168.
[15] B.V.S. Kumar, S.D. Vaidya, R.V. Kumar, S.B. Bhirud, R.B. Mane, Eur. J. Med.
Chem. 41 (2006) 599e604.
of structure-activity relationships, it could be noted that electron
donating groups like eNH2, 2-chloro and 2,4-dichloro on the phenyl
ring at 5th position of oxadiazole moiety have great influence on
anticancer activityand groups like eN(C2H5)2, eOeC6H5 on methyl at
5th position of oxadiazole ring also resulted in compounds with
improved anticancer activity. Electron withdrawing groups like
eCOOH and eC]O did not show remarkable anticancer activity. The
most active compound of the series was 1beA18 (2,4-dichloro
substituted) therefore, further studies on this compound continue in
our research laboratory to acquire more information about QSAR.
Finally it is conceivable that further derivatization of these
compounds could result in obtaining more selective anticancer
agents.
[16] K.M. Hosamani, H.R. Seetharamareddy, R.S. Keri, M.S. Hanamanthagouda,
M.G. Moloney, J. Enz. Inhi. Med. Chem. 24 (2009) 1095e1100.
[17] O.O. Guven, T. Erdogan, H. Goker, S. Yildiz, Bioorg. Med. Chem. Lett. 17 (2007)
2233e2236.
[18] H. Goker, C. Kus, D.W. Boykin, S. Yldz, N. Altanlar, Bioorg. Med. Chem. 10
(2002) 2589e2596.
[19] I. Kerimov, G.A. Kilcigil, B.C. Eke, N. Altanlar, J. Enz. Inhi. Med. Chem. 17 (2007)
696e701.
[20] G.A. Kilcgil, C. Kus, T. Coban, B.C. Eke, M. Iscan, J. Enz. Inhi. Med. Chem. 19
(2004) 129e135.
Acknowledgements
We are thankful to UGC, New Delhi, Government of India, for
financial assistance in the form of Major Research Project [36-107/
2008(SR)]. Thanks are also due to National Cancer Institute (NCI),
USA, for in vitro anticancer screening of the compounds.
[21] R. Franski, Asian J. Chem. 17 (4) (2005) 2063e2075.
[22] V. Padmavathi, S.G. Reddy, A. Padmaja, P. Kondaiah, A. Shazia, Eur. J. Med.
Chem. 44 (2009) 2106e2112.
[23] A.S. Aboraia, H.M. Abdel-Rahman, N.M. Mahfouz, M.A. El-Gendy, Bioorg. Med.
Chem. 14 (2006) 1236e1246.
[24] D. Kumar, G. Patel, E.O. Johnson, K. Shah, Bioorg. Med. Chem. Lett. 19 (2009)
2739e2741.
[25] P. Sengupta, D.K. Dash, V.C. Yeligar, K. Murugesh, D. Rajalingam, J. Singh,
T.K. Maity, Indian. J. Chem. 47B (2008) 460e462.
Supplementary data
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.ejmech.2012.04.027. These data
include MOL files and InChIKeys of the most important compounds
described in this article.
[26] D. Kumar, G. Sundaree, E.O. Johnson, K. Shah, Bioorg. Med. Chem. Lett. 19
(2009) 4492e4494.
References
[27] A.S. Kiselyov, M.N. Semenova, N.B. Chernyshova, A. Leitao, A.V. Samet,
K.A. Kislyi, M.M. Raihstat, T. Oprea, H. Lemcke, M. Lantow, D.G. Weiss,
N.N. Ikizalp, S.A. Kuznetsov, V.V. Semenov, Eur. J. Med. Chem. 45 (2010)
1683e1697.
[28] M.R. Boyd, K.D. Paull, Drug Dev. Res. 34 (1995) 91e109.
[29] J.N. Weinstein, T.G. Myers, P.M. O’Connor, S.H. Friend, A.J. Fornance Jr.,
K.W. Kohn, T. Fojo, S.E. Bates, L.V. Rubinstein, N.L. Anderson, J.K. Buolamwini,
W.W. van Osdol, A.P. Monks, D.A. Scudiero, E.A. Sausville, D.W. Zaharevitz,
B. Bunow, V.N. Viswanadhan, G.S. Johnson, R.E. Wittes, K.D. Paull, Science 275
(1997) 343e349.
[1] S.X. Cai, Recent Patents Anti-cancer Drug Discov. 2 (20070 79e101).
[2] L.M. Shi, Y. Fan, J.K. Lee, M. Waltham, D.T. Andrews, U. Scherf, K.D. Paull,
J.N. Weinstein, J. Chem. Inf. Comput. Sci. 40 (2000) 367e379.
[3] A. Monks, D. Scudiero, P. Skehan, R. Shoemaker, K. Paull, D. Vistica, C. Hose,
J. Langley, P. Cronise, V.A. Wolff, M.G. Goodrich, H. Campbell, J. Mayo,
M.J. Boyd, Natl. Cancer Inst. 83 (1991) 757e766.
[4] Y.H. Yang, M.S. Cheng, Q.H. Wang, H. Nie, N. Liao, J. Wang, H. Chen, Eur. J. Med.
Chem. 44 (2009) 1808e1812.
[5] A.G. Shadia, H. Khaled, S.K. Ahmed, L.R. Mireya, M.K. Sean, M.A. Abdel,
I.E. Hoda, Eur. J. Med. Chem. 44 (2009) 1500e1508.
[31] R.H. Shoemaker, Nat. Rev. Cancer 6 (2006) 813e823.