Amide derivatives of 4-azaindole: Design, synthesis, and EGFR targeting anticancer agents

Article abstract of DOI:10.1080/00397911.2019.1683206

A series of amide derivatives of azaindole-oxazoles (11a–n) were designed and synthesized and their structures were confirmed by ¹HNMR, ¹³CNMR and mass spectral analysis. Further, these derivatives were screened for their anticancer activity against human cancer cell lines viz; MCF7 (breast), A549 (lung) and A375 (melanoma). In vitro anticancer activity screening indicated that most of the hybrids exhibited potent inhibitory activities in a variety of cancer cell lines. Among the compounds 11d, 11e, 11f, 11j, 11k, 11l, 11m, and 11n were exhibited more potent activity than standard, in those mainly two compounds 11m and 11j were exhibited excellent activity in MCF-7 cell line with IC₅₀ values 0.034 and 0.036 μM. Moreover, all these compounds were carried out their molecular docking studies on EGFR receptor results indicated that two potent compounds 11m and 11j were strongly binds to protein EGFR (PDB ID: 4hjo). It was found that the energy calculations were in good agreement with the observed IC₅₀ values.

Full text of DOI:10.1080/00397911.2019.1683206

Synthetic Communications
An International Journal for Rapid Communication of Synthetic Organic
Chemistry
ISSN: 0039-7911 (Print) 1532-2432 (Online) Journal homepage: https://www.tandfonline.com/loi/lsyc20
Amide derivatives of 4-azaindole: Design,
synthesis, and EGFR targeting anticancer agents
Puli Venkat Swamy, Vukoti Kiran Kumar, Ruddarraju Radhakrishnam Raju,
Regalla Venkata Reddy, Anindita Chatterjee, Gangarapu Kiran & Gattu
Sridhar
To cite this article: Puli Venkat Swamy, Vukoti Kiran Kumar, Ruddarraju Radhakrishnam Raju,
Regalla Venkata Reddy, Anindita Chatterjee, Gangarapu Kiran & Gattu Sridhar (2019): Amide
derivatives of 4-azaindole: Design, synthesis, and EGFR targeting anticancer agents, Synthetic
Communications, DOI: 10.1080/00397911.2019.1683206
To link to this article: https://doi.org/10.1080/00397911.2019.1683206
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https://doi.org/10.1080/00397911.2019.1683206
Amide derivatives of 4-azaindole: Design, synthesis, and
EGFR targeting anticancer agents
Puli Venkat Swamya, Vukoti Kiran Kumara, Ruddarraju Radhakrishnam Rajub,
Regalla Venkata Reddya, Anindita Chatterjeea, Gangarapu Kiranc, and
Gattu Sridhard
aDepartment of Chemistry, Koneru Lakshmaiah Education Foundation, Guntur, Andhra Pradesh, India;
c
bDr. MACS Bio-Pharma Pvt. Ltd, Medak Telangana, India; Department of Pharmaceutical Analysis and
d
Chemistry, School of Pharmacy, Anurag Group of Institutions, Medchal, Telangana, India; Organic and
Biomolecular Chemistry Division, CSIR-Indian Institute of Chemical Technology, Hyderabad, India
ABSTRACT
ARTICLE HISTORY
Received 14 July 2019
A series of amide derivatives of azaindole-oxazoles (11a–n) were
designed and synthesized and their structures were confirmed by
1HNMR, 13CNMR and mass spectral analysis. Further, these derivatives
were screened for their anticancer activity against human cancer cell
lines viz; MCF7 (breast), A549 (lung) and A375 (melanoma). In vitro
anticancer activity screening indicated that most of the hybrids
exhibited potent inhibitory activities in a variety of cancer cell lines.
Among the compounds 11d, 11e, 11f, 11j, 11k, 11l, 11m, and 11n
were exhibited more potent activity than standard, in those mainly
two compounds 11m and 11j were exhibited excellent activity in
MCF-7 cell line with IC50 values 0.034 and 0.036 mM. Moreover, all
these compounds were carried out their molecular docking studies
on EGFR receptor results indicated that two potent compounds 11m
and 11j were strongly binds to protein EGFR (PDB ID: 4hjo). It was
found that the energy calculations were in good agreement with the
observed IC50 values.
KEYWORDS
Azaindole; oxazole; docking
and anticancer activity
GRAPHICAL ABSTRACT
O
N
R
H
O
N
R = H, 4-Me, 3,5-diMe, 4-OMe,
3,5-diOMe, 3,4,5-triOMe, 4-Br, 3,5-diBr,
4-Cl, 3,5-diCl, 4-F, 4-NO2,
3,5-diNO2 ,4-CN
N
N
N
H
N
H
Introduction
Nitrogen-containing heterocyclic compounds play a very important role in medicinal
and pharmaceutical chemistry. Indole is a nitrogen-containing fused heterocyclic
CONTACT Puli Venkat Swamy
venkatswamichem1@gmail.com
Department of Chemistry, Koneru Lakshmaiah
Education Foundation, Vaddeswaram, Guntur, Andhra Pradesh, 522502, India.
Color versions of one or more of the figures in the article can be found online at www.tandfonline.com/lsyc.
Supplemental data for this article can be accessed on the publisher’s website.
ß 2019 Taylor & Francis Group, LLC

2
P. VENKAT SWAMY ET AL.
Figure 1. Anticancer target indole molecules (I & II) and biologically active indole-oxazole mol-
ecule (III).
compound and widely distributed in many bioactive compounds and natural products[1]
such as alkaloids, plant, animal and microbial hormones. Among the based on indole
drugs, Indiublin and LKBI showed anticancer activity against various cancer cell lines.
Indibulin (I, Figure 1) (also called ZIO-301 and D-24851) is a novel synthetic anti-
mitotic agent that binds tubulin, destabilizes microtubulin polymerization and arrests
tumor cell growth at the G2/M phase. Microtubulins are well-established targets for
anti-cancer drug development[2,3] and also Indibulin had been used Phase I/II trials for
the treatment of metastatic breast cancer (NCT01113970) and Liver kinase B1 (II,
Figure 1) (LKB1, also known as STK11) was first identified as the causal mutation in
Peutz–Jeghers Syndrome (PJS), a rare inherited autosomal dominant disorder and it was
characterized by the development of gentle gastrointestinal hamartomas and the early
onset of cancer.[4] However, more than a decade later, LKB1 has become recognized as
a critical tumor-suppressor gene that is regularly mutated in a broad spectrum of
human cancers. Similarly, 4-azaindoles have generated an increased interest due to their
structural similarities with indoles and played key role in medicinal chemistry.[5–7]
These were exhibited a variety of biological activities such as antimalarial,[8] anthelmin-
tic,[9] non-narcotic antitussive,[10] anti-diabetic,[11] antiviral,[12] anticholinesterase,[13,14]
and anticancer.[15–17]
During the last century, much research has concentrated on the isolation, biological
activities and synthesis of indole-oxazole due to their biological and pharmaceutical sig-
nificance. For example, pimprinine (III, Fig. 1) is an indole-alkaloid and demonstrated
good monoamine oxidase (MAO) inhibition[18] and antiepileptic[19] activity. In addition,
oxazoles are five-member heterocyclic essential structural units for medicinal chemists
and have attracted continuing interest over the years. Which are displayed versatile

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SYNTHETIC COMMUNICATIONSV
3
OH
N
CHO
CN
N
N
N
N
Ac2O
NH2OH.HCl
HMTA
AcOH, H2O
reflux, 6 h, 79%
140 °C, 1 h, 89%
N
Pyridine
115 °C, 6 h, 87%
N
N
H
N
H
H
H
3
5
4
1
EtOH
80 °C, 4 h,
88%
Aq.NaOH
O
O
O
OEt
OH
7
O
O
O
N
NH2
N
Br
O
N
N
O
N
Aq.NaOH
EtOH
N
H
EtOH
80 °C, 5 h, 85%
N
H
N
rt, 10 h, 86%
H
6
8
9
NH2
EDCI, HOBt
10a
; R = H
10b; R = 4-Me
10c; R = 3,5-diMe
10d;
11a
; R = H
DCM
rt, 12h,
R 10a-n
11b; R = 4-Me
11c
; R = 3,5-diMe
R = 4-OMe
10e; R = 3,5-diOMe
11d; R = 4-OMe
11e; R = 3,5-diOMe
O
10f
; R = 3,4,5-triOMe
11f
; R = 3,4,5-triOMe
11g; R = 4-Br
11h
10g; R = 4-Br
10h; R = 3,5-diBr
N
H
R
; R = 3,5-diBr
O
10i
; R = 4-Cl
10j; R = 3,5-diCl
10k;
N
11i; R = 4-Cl
11j; R = 3,5-diCl
N
R = 4-F
11k;
R = 4-F
11l; R = 4-NO2
11m;
10l; R = 4-NO2
10m;R = 3,5-diNO2
10n
11a-n
(69% - 92%)
N
H
R = 3,5-diNO2
11n; R = 4-CN
; R = 4-CN
Scheme 1. 4-Azaindole amide derivatives.
biological activities including anticancer,[20] A2A adenosine receptor antagonists,[21] anti-
HIV,[22] anti-tuberculotic,[23] antifungal,[24] insecticidal,[25] herbicidal[26] and receptor
tyrosine kinases (RTK).[27]
Based on the biological evaluation and mechanism studies of anticancer indoles
revealed that these compounds have been showing target diverse pathways in cancer
cells. Accordingly, biological importance and current application of indole-derived com-
pounds (I, II, & III) as shown in (Figure 1), we have designed and synthesized novel
amide derivatives of azaindole-oxazole as anticancer agents. This manuscript summa-
rizes the versatility of indole-oxazole core structure and SAR studies, arranged by their
biological activity, which leads to the discovery of diverse new anticancer agents and
subsequently in silico molecular docking were evaluated.
Results and discussion
Chemistry
The synthetic route of target compounds (11a-n) as shown in Scheme 1. Compound 1
was treated with HMTA in acetic acid and H2O at reflux for 6 hours to afford pure
aldehyde compound 3 in good yield. Next, the carbonyl addition by compound 3 with

4
P. VENKAT SWAMY ET AL.
Table 1. In vitro cytotoxic activity of compounds (11a–n) in (IC50 mM).
Comp No
R
MCF-7 (breast)
A-549 (Lung)
A375 (Melanoma)
11a
11b
11c
11d
11e
11f
11g
11h
11i
11j
11k
H
4-Me
3,5-diMe
4-OMe
3,5-diOMe
3,4,5-triOMe
4-Br
3,5-diBr
4-Cl
3,5-diCl
4-F
4-NO2
3.78 0.31
3.89 0.29
14.39 0.99
1.11 0.12
2.40 0.17
0.90 0.11
6.10 0.49
3.76 0.31
4.89 0.36
0.036 0.0023
0.66 0.051
0.22 0.019
0.034 0.002
1.23 0.093
2.02 0.17
–
–
4.34 0.39
7.32 0.66
0.78 0.065
1.73 0.14
1.56 0.13
–
0.95 0.083
8.67 0.71
0.92 0.081
1.33 0.11
0.14 0.013
0.59 0.044
0.28 0.022
2.18 0.18
9.67 0.91
5.30 0.39
1.54 0.13
0.84 0.069
2.67 0.19
–
1.25 0.093
15.9 1.1
1.74 0.13
–
11l
–
11m
11n
Doxorubicin
3,5-diNO2
4-CN
0.28 0.019
1.77 0.14
5.51 0.039
The results were expressed as the (IC50 mM). “–” ¼ Not active.
Values are mean SEM.
hydroxylamine hydrochloride in pyridine at 115 ꢀC for 6 hours to afford pure oxime for-
mation compound 4. Then, nitrile formation by reacted with acetic anhydride at 140 ꢀC
for 1 hour to afford pure cyano compound 5 and then followed by partial hydrolysis
with aqueous NaOH in ethanol at 80 ꢀC for 4 hours to afford amide compound 6. This
amide intermediate was reacted with ethyl 3-bromo-2-oxopropanoate 7 in ethanol at
reflux for 5 hours to give oxazole compound 8. Afterward, this intermediate 8 under-
goes basic hydrolysis with aqueous NaOH in ethanol at room temperature for 10 hours
to give acid compound 9. Finally, this acid 9 was coupled with different substituted aro-
matic amines (10a–n) by using reagents EDCI, HOBt in CH2Cl2 at room temperature
for 12 hours to afford target compounds (11a–n). The structures of the synthesized
1
compounds were confirmed by H NMR, 13CNMR and mass spectral data. As a repre-
sentative example, the 1HNMR spectra of N-(3,5-dinitrophenyl)-2-(1H-pyrrolo[3,2-
b]pyridin-3-yl)oxazole-4-carboxamide (11 m) is characterized as follows: One broad
singlet appears at 9.57 (brs, 1 H) ppm corresponds to amide proton –CONH–, one sing-
let at 9.73 (s, 2 H) corresponds to –ArH and one singlet appear at 9.87 (s, 1 H) corre-
sponds to isoxazole –H. 13C NMR spectral data shown singlet peak appear at 145.8 ppm
is corresponds to isoxazole –C–, singlet at 159.7 ppm corresponds to isoxazole
–O–C¼N and singlet appear at 161.8 ppm corresponds to keto carbon –CO–. The mass
spectra of the synthesized compounds showed 395 [M þ H]þ peaks in agreement with
their molecular formula.
Biological evaluation
In vitro cytotoxicity
Newly synthesized compounds (11a–n) were tested for their anticancer activity against
three human cancer cell lines including breast (MCF7), lung (A549) and melanoma
(A375) by MTT assay. Here positive control used as doxorubicin and results are
expressed in IC50 with mM and incorporated in Table 1 and also shown in bar diagram
Figure 2. In vitro anticancer activity, screening indicated that most of the hybrids exhib-
ited potent inhibitory activities in a variety of cancer cell lines with low micromolar to

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5
Figure 2. Docking pose of compound 11j (Ball and stick model) in the EGFR (PDB ID: 4hjo) binding
pocket. White line represents the hydrogen bonding. Only the key residues are shown for clarity.
submicromolar IC50 values. Among them, compounds 11d, 11e, 11f, 11j, 11k, 11 l,
11 m and 11n were exhibited more potent activity than standard. Especially two com-
pounds 11 m and 11j were exhibited excellent activity against MCF-7 cell line with IC50
values 0.034 and 0.036 mM. In general as compared with the number of cell wises, the
highest activity shown by the compounds 11 m and 11j were the most active and effect-
ive against three cell lines, in which compounds 11 m (IC50 ¼ 0.034 0.002, 0.59 0.044,
and 0.28 0.019 mM) and 11j (IC50 ¼ 0.036 0.0023, 0.92 0.081 and 1.74 0.013 mM)
for MCF7 (Breast), A549 (Lung) and A375 (Melanoma), respectively. The next better
activity showed by compounds 11 l and 11k against two cancer cell lines, in which com-
pound 11 l (IC50 ¼ 0.22 0.019 and 0.14 0.013 mM) and 11k (IC50¼0.66 0.051 and
1.33 0.11 mM) for MCF7 (Breast) and A549 (Lung), respectively. The remaining com-
pounds 11d, 11e, 11f, and 11n also showed good inhibition of three or two cancer cell
lines such as breast, lung, and melanoma as shown in Table 1.
The structure-activity relationships (SARs) were established from the anticancer data
reported in Table 1. Firstly, the electron-donating analogs (11a-f) were described by the
most active compound 11d at the para position of 4-methoxy showed more potent activity.
Further, in this way introduced more methoxy groups at phenyl ring part by tri
methoxy analog 11f at di meta and para positions and compound 11e at di meta posi-
tions results were observed decreasing activity. In that next, simple phenyl analog 11a
and methyl substituted analogs 11b and 11c were not active as compared with methoxy
analogs (11d–f) and also with the standard drug.

6
P. VENKAT SWAMY ET AL.
Secondly, in the electron with-drawing series (11g–n) a dramatic change in activity
was observed. The most active analogs such as dinitro 11m and dichloro 11j at meta
positions of phenyl ring displayed activity in micro molars. In such a way, the next bet-
ter activity compounds 11l >11k introduced nitro > fluoro at para position displayed
loss in the melanoma cell. Consequently, compound 11n at the para position of cyano
resulted in good activity as compared with halogen analogs 11h, 11i and 11 g displayed
weaker activities. Finally, in conclusion, most of the with-drawing substitution analogs
(11j–n) at the phenyl part were found to be the most active of the series. In summary,
the information of SAR provided us a guideline to improve the anticancer activity in
future structural modification.
Molecular docking studies
The protein receptors EGFR is a striking target in the cancer disease. The epidermal
growth factor receptor (EGFR) is the cell-surface receptor for members of the epidermal
growth factor family of extracellular protein ligands.[28] It lives on the cell surface and
driven by binding of its specific ligands it plays a major role in the ductal development
of the mammary glands.[29–31] Overexpression of EGFR associated with a number of
cancers, including squamous-cell carcinoma of the breast, lung, anal cancers,[32] glio-
blastoma and epithelian tumors of the head and neck.[33] These somatic mutations
involving EGFR lead to its constant activation, which produces uncontrolled
cell division.
In this regard, we have screened all compounds in molecular docking studies and we
found interesting idea in the process of exploration of results. In that two molecules 11j
and 11 m approaches strongly to receptor EGFR protein as showing by their minimum
binding energies 68.85 and 68.58 Kcal/mol, respectively. The docking results were found
to be in good agreement with in vitro experimental IC50 values (Table 2). Figures 3 and 4
show the best conformations of 11j and 11 m forming a cluster into the binding pocket
of EGFR receptor. The GOLD program was used to identify favorable conformations and
possible interaction regions at the active site of the EGFR enzyme (4HJO). All the top-
ranked conformations of (11a–n), were orientated in such a way as to insert themselves
Table 2. Docking interations.
Binding Energy
Number of
hydrogen bonds
S.No.
Compound
(kcal mol–1
)
Residues involved in hydrogen bonding
1
2
3
4
5
6
7
8
11a
11b
11c
11d
11e
11f
11g
11h
11i
11j
11k
11l
11m
11n
AQ4
62.59
64.05
65.41
66.52
66.06
68.20
62.75
66.46
62.82
68.85
60.22
60.23
68.58
62.47
71.91
3
02
03
03
02
03
03
03
01
3
2
3
3
02
03
Asp831(3.06), Met769(2.60), Thr830(2.64)
Thr766 (2.56), Met769(2.28)
Asp831(2.91), Thr766(2.59), Met769 (2.59)
Met769(2.69),Thr830(2.60).Asp831(2.88)
Met769(3.06), Thr766 (2.97)
Thr830(2.05), Asp831 (2.96), Lys721(2.72)
Asp831(2.98), Thr766(2.64), Met769(2.42)
Asp831(2.99), Thr766 (2.76), Met769(2.47)
Met769(2.74)
Thr766(2.50), Asp831(2.90), Met769(2.55)
Met769(2.54), Thr766(2.57)
9
10
11
12
13
14
15
Met769(2.56), Asp831(2.72), Thr830(2.76)
Met769(2.72), Asp831(2.75), Thr830(2.79)
Leu764(2.88) Met769(2.86)
Thr766(2.34),Asp831(2.42), Met769(2.57)

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7
into the same pocket of the EGFR structure, and their docked locations on the active
site were very close to that occupied by the co-crystallized ligand ‘AQ4’. Figure 5
shows a superimposition of these docking solutions alongside the docking of the lig-
and ‘11j’ that was co-crystallized with EGFR. In an attempt to explain the differences
between the docking scores, an analysis of the polar interactions (hydrogen bonds) for
the docked ligands on the active site of EGFR structure are summarized in Table 2
(in supplementary material) along with the distances between the donor atoms
involved in this interaction by way of ligand–enzyme complexation. Additionally,
details were provided of the docking positions for a number of these derivatives
(Figures 3–5). Docking positions were captured using the Hermes program.
The compound 11j, which possesses a chloro substituent, which presented the most
stable GOLD score. By contrast, another nitro analog 11m also presented a stable
GOLD score. Figure 3 shows the analog 11j establishing H-bonds with the Thr766,
Asp831, Met769 residues, with distances of 2.50, 2.90 and 2.55 Å, respectively.
Likewise, Figure 3 demonstrates that Asp831 is involved in H-bond, the nitrogen
group of azaindole ring, Thr766 forms H-bond with oxygen group of oxazole and Met
769 with oxygen group of amide. While Figure 4 shows that the 11 m derivative forms
H-bonds with Met769, Asp831, and Thr830 of 2.72, 2.75, and 2.79 Å in length,
respectively.
Figure 3. Docking pose of compound 11 m (Ball and stick model) in the EGFR (PDB ID: 4hjo) binding
pocket. White line represents the hydrogen bonding. Only the key residues are shown for clarity.

8
P. VENKAT SWAMY ET AL.
Figure 4. Docked pose of compounds 11j (Blue, ball and stick) and alongside the cocrystallized ligand
‘AQ4’ (yellow, ball and stick model) in the EGFR binding pocket.
Likewise, Figure 4 demonstrates that Asp831 is involved in the H-bond with the
nitrogen group of azaindole ring, Met769 forms H-bond with nitro group on benzene
and Thr830 with nitro group of oxazole.
All these derivatives (11a–n) have shown constant H-bond interactions with amino
acids Met769, Asp831, Thr830. In conclusion, the experimental in vitro results strongly
correlate with the molecular docking analysis.
Furthermore, the docking has been validated by using Auto dock and the compounds
11j and 11 m have shown same interactions with EGFR (4HJO). The compound 11j has
shown (Figure 5) H-bond interaction Met769 and Asp 831 and similarly, 11 m has
shown (Figure 6) H-bond interactions with Met769, Lys721, Arg752, and Thr766.
Drug-likeness and ADME properties
Bioavailability of amide derivatives of 4-azaindole (11a–n) was accessed through ADME
(Adsorption, Distribution, Metabolism, and Excretion) using molinspiration. In order to
explore drug-like properties of compounds (11a–n), the lipophilicity, expressed as the
octanol/water partition coefficient and here it is called logP(o/w), as well as other theor-
etical calculations such as molecular size, the number of hydrogen bond acceptors and
donors, TPSA and % ABS: Percentage of absorption as shown in Table 3.

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9
Figure 5. Docking pose of compound 11j in the EGFR (PDB ID: 4hjo) binding pocket.
The violation of more than one of these rules may indicate problems in the bioavail-
ability of the potential drugs. The results showed all the compounds complied with
Lipinski’s rule and meet the criteria as shown in Table 3.
Toxicity risk assessment screening
The toxic properties such as mutagenic, tumorigenic, irritant and reproductive effects
were screened for the 4-azaindole amide derivatives (11a–n) using Molinsperation ser-
ver. The server is inbuilt with list of about 5300 distinct substructure fragments created
by 15,000 commercially available fragments with reported drug score and drug-likeness.
Drug score is associated with clogP, drug likeness and toxicity risks as a total value may
be used to judge the overall potential to qualify it as a drug. The toxicity screening of
all compounds (11a–n) showed that no risk of mutagenic, tumorigenic, irritant and
reproductive toxicity as shown in Table 4.
Experimental section
All chemicals and reagents were obtained from Aldrich (Sigma–Aldrich, St. Louis, MO,
USA), Lancaster (Alfa Aesar, Johnson Matthey Company, Ward Hill, MA, USA) and
were used without further purification. Reactions were monitored by TLC, performed
on silica gel glass plates containing 60 F-254, and visualization on TLC was achieved by
UV light or iodine indicator. 1H and 13C NMR spectra were recorded on Bruker,

10
P. VENKAT SWAMY ET AL.
Figure 6. Docking pose of compound 11 m in the EGFR (PDB ID: 4hjo) binding pocket.
Table 3. ADME properties.
Compound Rule
logP(o/w)<5
MW<500
TPSA<140
HBA<10
HBD<5
nRotB<10
Volume
11a
11b
11c
11d
11e
11f
11g
11h
11i
11j
11k
11l
11m
11n
2.53
2.97
3.35
2.58
2.57
2.16
3.33
4.07
3.20
3.81
2.69
2.48
2.37
2.28
304.09
318.34
332.36
334.33
364.36
394.39
383.20
462.10
338.75
373.20
322.30
349.31
394.30
329.32
83.81
83.81
83.81
93.05
102.28
111.52
83.81
83.81
83.81
83.81
83.81
129.64
175.46
107.61
6
6
6
7
8
9
6
6
6
6
6
9
12
7
2
2
2
2
2
2
2
2
2
2
2
2
2
2
3
3
3
4
5
6
3
3
3
3
3
4
5
3
260.48
277.06
293.60
286.02
311.57
337.12
278.37
296.25
274.01
287.55
265.41
283.8
307.15
277.34
Bruker UXNMR/XWIN-NMR (400 MHz, 300 MHz) instrument. Chemical shifts (d) are
reported in ppm downfield from internal TMS standard. ESI spectra were recorded on
Micro mass, Quattro LC using ESI þ software with capillary voltage 3.98 kV and ESI
mode positive ion trap detector. Melting points were determined with an electrothermal
melting point apparatus and are uncorrected.
General procedure for the synthesis of compounds (11a–n):
All the final reactions (11a–n) were performed by 200 mg scale. To a stirred solution of
acid compound 9 (1.0 equiv) and substituted aromatic amines (10a–n) (1.0 equiv) in

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11
Table 4. Toxicity risk assessment of compounds (11a–n).
Compound cLogP Solubility Drug likeness Drugscore Mutagenic Tumorigenic Irritant Reproductive effect
11a
11b
11c
11d
11e
11f
11g
11h
11i
11j
11k
11l
11m
11n
2.24
2.58
2.93
2.17
2.1
2.03
2.93
3.69
2.84
3.45
2.34
1.32
0.39
2.07
ꢂ4.37
ꢂ4.71
ꢂ5.06
ꢂ4.39
ꢂ4.41
ꢂ4.42
ꢂ5.2
1.13
1.46
0.67
0.65
0.39
0.69
0.37
0.71
0.32
0.24
0.65
0.34
0.66
0.35
0.32
0.34
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
No
ꢂ1.57
1.69
No
No
ꢂ6.44
4.33
0.89
Medium
Medium
Medium
Medium
Medium
Medium
No
No
No
No
Medium
Medium
Medium
Medium
Medium
Medium
No
No
No
No
ꢂ6.04
ꢂ5.11
ꢂ5.84
ꢂ4.68
ꢂ4.83
ꢂ5.29
ꢂ5.14
ꢂ2.94
3.75
ꢂ1.08
1.64
ꢂ12.5
ꢂ7.86
ꢂ3.84
dichloromethane was added EDCI (1.0 equiv), HOBt (1.0 equiv) and DIPEA (1.2 equiv)
at room temperature. The reaction mixture was stirred for 12 h. After completion of the
reaction as monitored by TLC, the reaction mixture was diluted with dichloromethane,
washed with saturated solution of NaHCO3, dried over anhydrous Na2SO4, filtered and
concentrated. The crude products were purified by column chromatography eluted with
ethyl acetate/hexane to afford the pure compounds (11a–n).
N-Phenyl-2-(1H-pyrrolo[3,2-b]pyridin-3-yl)oxazole-4-carboxamide (11a)
The general procedure 11a–n was followed for the synthesis of 11a, 211.3 mg in 80% yield.
1
Mp:196–198 ꢀC, H NMR (400MHz, DMSO-d6): d 7.30–7.41 (m, 4H), 7.48–7.52 (m, 2H),
7.71 (d, 2H, J ¼ 7.23Hz), 8.89 (d, 1H, J ¼ 8.19Hz), 9.24 (s, 1H), 9.56 (brs, 1H), 9.86 (s, 1H);
13C NMR (75MHz, DMSO-d6): d 97.4, 116.4, 118.6, 122.4, 124.5, 124.9, 131.4, 138.4, 138.9,
139.3, 141.2, 144.8, 145.8, 158.3, 159.8; HRMS (ESI): m/z calculated for C17H12N4O2Na
[Mþ Na]þ 327.0858, found 327.0851.
Anticancer assay
MTT protocol
The cytotoxic activity of the compounds was determined using the MTT assay. 1 ꢁ 104
cells/well were seeded in 200 mL DMEM, supplemented with 10% FBS in each well of
96-well microculture plates and incubated for 24 hours at 37 ꢀC in a CO2 incubator.
Compounds, diluted to the desired concentrations in culture medium, were added to
the wells with respective vehicle control. After 48 hours of incubation, 10 mL MTT (3-
(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) (5 mg/mL) was added to
each well and the plates were further incubated for 4 hours. Then the supernatant from
each well was carefully removed, formazon crystals were dissolved in 100 mL of DMSO
and absorbance at 540 nm wavelength was recorded.

12
P. VENKAT SWAMY ET AL.
Docking assay
Molecular docking protocol
Gold 5.6 tool was employed for the in silico docking studies to explore the binding
mode of ligands and were docked into cavity EGFR (PDB ID: 4HJO). The binding pro-
file for azaindole-oxazole amide derivatives (11a–n) ligands with 4HJO was determined
and for each ligand ten conformations were generated. The conformations which are
having highest ranking were used for further analysis and Gold fitness calculated using
Gold docking function.
Conclusion
In conclusion, we have designed and synthesized a novel series of amide derivatives of
azaindole oxazoles (11a–n) and were evaluated anticancer activity against three human
cancer cell lines. Most of the compounds showed moderate to excellent anticancer activ-
ity, in those eight analogs 11d, 11e, 11f, 11j, 11k, 11l, 11m, and 11n were showed
more potent activity than compared with reference drug. The highest activities were
found for compounds 11m and 11j with IC50 values of 0.034 and 00.36 mM in
MCF7cell, respectively. Moreover, compound 11m and 11j proved to be 58 to 60-folds
more active in MCF7 cell as compared with standard. Moreover, molecular docking
studies on EGFR receptor results indicated that two compounds 11m and 11j of bind-
ing interactions strongly correlated with in vitro anticancer activity. Further changes
and structural modifications of these analogs in order to raise their potency is currently
in progress.
1
Full experimental details, spectral data of the products, H NMR and 13C NMR of all
the new compounds can be found via the Supplementary Content section of this
article’s Web page.
Acknowledgments
We would like to express our gratitude and thanks to the Department of Chemistry, Koneru
Lakshmaiah University, Guntur for constant encouragement and providing basic research facility.
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