Synthesis and biological evaluation of phenyl-amino-pyrimidine and indole/oxindole conjugates as potential BCR-ABL inhibitors

Article abstract of DOI:10.1007/s00044-019-02318-4

Abstract: Indole/isatin conjugated phenyl-amino-pyrimidine derivatives have been synthesized, characterized and evaluated in vitro for their potential as BCR-ABL inhibitors. Among the series, all derivatives (7a–7o) were found to be more cytotoxic than standard Imatinib against K-562 cell line. Compound 7l was the most active in the series with almost two folds more potency than imitanib (IC₅₀ 0.65 μM). In vitro enzymatic studies with recombinant ABL kinase enzyme exhibited promising inhibition in the range of 30–71 μM for most of these novel conjugates. In addition, modelling and other computational studies have been carried out to draw insight into the BCR-ABL protein interactions with the target molecules and drug like properties of the conjugates, respectively. [Figure not available: see fulltext.].

Full text of DOI:10.1007/s00044-019-02318-4

MEDICINAL
CHEMISTRY
RESEARCH
Medicinal Chemistry Research
https://doi.org/10.1007/s00044-019-02318-4
ORIGINAL RESEARCH
Synthesis and biological evaluation of phenyl-amino-pyrimidine and
indole/oxindole conjugates as potential BCR-ABL inhibitors
Abdul Rahim1,2 Riyaz Syed
1,3 ●
Y. Poornachandra M. Shaheer Malik Ch. Venkata Ramana Reddy
4 ●
5 ●
3 ●
●
Mallika Alvala6 Kiran Boppana B. Sridhar Ramars Amanchy Ahmed Kamal
7 ●
8 ●
4 ●
1,6,9
●
Received: 22 October 2018 / Accepted: 16 February 2019
©
Springer Science+Business Media, LLC, part of Springer Nature 2019
Abstract
Indole/isatin conjugated phenyl-amino-pyrimidine derivatives have been synthesized, characterized and evaluated in vitro
for their potential as BCR-ABL inhibitors. Among the series, all derivatives (7a–7o) were found to be more cytotoxic than
standard Imatinib against K-562 cell line. Compound 7l was the most active in the series with almost two folds more potency
than imitanib (IC50 0.65 μM). In vitro enzymatic studies with recombinant ABL kinase enzyme exhibited promising
inhibition in the range of 30–71 µM for most of these novel conjugates. In addition, modelling and other computational
studies have been carried out to draw insight into the BCR-ABL protein interactions with the target molecules and drug like
properties of the conjugates, respectively.
Graphical Abstract
Synthesis and biological evaluation of phenyl-amino-pyrimidine and indole/oxindole conjugates as potential BCR-ABL
inhibitors.
These authors contributed equally: Abdul Rahim, Riyaz Syed
Supplementary information The online version of this article (https://
doi.org/10.1007/s00044-019-02318-4) contains supplementary
material, which is available to authorized users.
5
*
Ahmed Kamal
abdulrahimakot@gmail.com
Central Research Laboratory and Department of Chemistry,
Faculty of Applied Sciences, Umm Al-Qura University, 21955
Makkah, Saudi Arabia
1
Medicinal Chemistry and Biotechnology Division, CSIR-Indian
Institute of Chemical Technology (IICT), Hyderabad 500 007,
India
6
Department of Medicinal Chemistry, National Institute of
pharmaceutical Education and Research (NIPER),
Balanagar, Hyderabad 500 037, India
2
3
4
Academy of Scientific and Innovative Research, New Delhi 110
7
8
9
GVK Bio Sciences Pvt. Ltd., Plot No. 79, IDA, Mallapur,
Hyderabad 500076, India
0
25, India
Department of Chemistry, J.N.T. University,
Kukatpally, Hyderabad 500 085, India
X-ray Crystallography Division, CSIR-Indian Institute of
Chemical Technology, Hyderabad 500007, India
Division of Applied Biology, CSIR-Indian Institute of Chemical
Technology, Tarnaka, Hyderabad 500 085, India
School of Pharmaceutical Education and Research, Jamia
Hamdard, New Delhi 110062, India

Medicinal Chemistry Research
●
●
●
●
Keywords Phenyl-amino-pyrimidine Indole Oxindole Bcr-Abl inhibitors Chronic myeloid leukemia
Introduction
and solid tumors (Second-line therapy). However, the
common adverse effects of these marketed drugs include
skin rash, nausea, vomiting, and vascular adverse events
(VAEs) often leading to severe organ damage (Amala et al.
2013; Płużański and Piórek 2016; Valent 2011; Oren et al.
2015; Haguet et al. 2017; Herrmann 2016; Jason 2017;
Ateyya et al. 2017; Sodergren et al. 2014). Furthermore, the
third generation BCR-ABL inhibitors ponatinib is asso-
ciated with certain arterial occlusive events (Dahlen et al.
2016; Jonathan et al. 2016; Chatree et al. 2016; Gusarova
and Turkina 2016) and being approved by FDA with a
black box warning.
Hence, there is a need to develop novel BCR-ABL
inhibitors with enhanced drug safety and efficacy. Design of
imatinib based novel BCR-ABL inhibitors offers opportu-
nity of simpler, safer, and affordable therapeutics with lesser
side effects and possessing unique inhibitory mechanisms
which could address the issue of drug resistance.
On the other hand, indole and isatin are promising
moieties in the design of anti-cancer agents and they are
present in different anticancer drugs (Sidhu et al. 2015; El
Sayed et al. 2015; Prakash et al. 2018; Vine et al. 2009,
2013; Havrylyuk et al. 2011; Teng et al. 2016). Keeping
this in view, we envisaged hybridizing the promising phe-
nylamino pyrimidine and indole/oxindole scaffolds into a
single chemical entity and explore the biological effect
exerted by these conjugates as newer BCR-ABL inhibitors.
In our investigative strategy the amide moeity present in
imatinib is substituted with isatin/indole ring through a
bridging imine bond to afford new series of phenyl-amino-
pyrimidine and indole/oxindole conjugates. This was fol-
lowed by the evaluation of the therapeutic potency of these
conjugates against K-562 cancer cell line and recombinant
ABL kinase enzyme with comparison to standard drug
Tyrosine kinases (TKs) are the enzymes responsible to
initiate transfer of phosphate group from ATP to tyrosine, a
critical process in cellular machinery, and inhibition of TKs
is currently considered as one of the most significant
molecular targets in cancer drug discovery programs (Sta-
quinini et al. 2015; Wilhelm et al. 2004). Chronic myeloid
leukemia (CML) is a myelo-proliferative disorder asso-
ciated with the philadelphia chromosome, resulting in the
BCR-ABL fusion gene. This genetic abnormality results in
the formation of a unique gene product (BCR-ABL), which
is a constitutively active TK that is implicated in the
development of CML (Salesse and Verfaillie 2002; An et al.
2
010; Goldman and Melo 2008; Rebecca et al. 2018;
Zámečníkova 2010; Carter et al. 2016). The phenylamino
pyrimidine scaffold based BCR-ABL inhibitor, imatinib
drug is considered as a “gold standard” in the treatment of
patients with newly diagnosed CML (Fig. 1) (Karl Peggs
and Stephen Mackinnon 2003; Nida and Naveed 2014;
Druker 2003). However, the emergence of resistance
against imatinib in association with BCR/ABL1 mutations
(
2
Sawyers et al. 2002; Talpaz et al. 2002; Azevedo et al.
017; Valent 2007; O’Hare et al. 2007; Jabbour et al. 2009)
has become a challenge. Further second generation BCR-
ABL inhibitors such as nilotinib (Fig. 1), dasatinib, and
ponatinib have been developed for treating the CML
patients (Cortes et al. 2011, 2016; Hochhaus et al. 2008,
2
016; Milojkovic et al. 2012; Porkka et al. 2008; Hughes
et al. 2009; Kantarjian et al. 2007; Shah et al. 2014; Giles
et al. 2013; Gambacorti-Passerini et al. 2014; Kantarjian
et al. 2014; Nicolini et al. 2017; Lipton et al. 2016).
Similarly, another phenylamino pyrimidine derivative
NRC-AN-019 is in phase-II clinical trials for treating CML
Fig. 1 BCR-ABL inhibitors and
Indole/oxindole based inhibitors

Medicinal Chemistry Research
Scheme 1 Synthesis of
phenylamino pyrimidine
intermediate 6
Table 1 Optimization of the reaction conditions
o
b
Entry
Solvent
Catalyst
Temp ( C)
Time (h)
Yield (%)
a
1
2
3
4
5
6
7
H2O
–
30
40
30
60
60
80
40
16
8
10
30
55
85
90
75
44
H2O
AcOH
AcOH
AcOH
MeOH
MeOH
MeOH
EtOH
DCM
8
4
a
–
4
AcOH
AcOH
8
8
aNo catalyst used
bIsolated yield
imatinib. Furthermore, we studied the BCR-ABL protein
binding interactions and drug like properties of these newly
synthesized ligands.
order to achieve this, reaction between compound 6 and
indole-3-carboxaldehyde was first carried out in water at
room temperature and 40 °C, the progress of the reaction
was monitored by TLC, however the starting materials were
not completely consumed even until 16 h which is in turn
affected the product yield (Table 1, entry 1). Therefore, a
focused optimization study was carried out with different
solvents in the presence and absence of acetic acid as cat-
alyst at varying temperature. It was observed that the use of
catalytic amount of acetic acid afforded better yields of the
product in water (Table 1, entry 2). On switching from
water to methanol in the presence of catalytic acetic acid at
room temperature the yield of the reaction improved to 55%
(Table 1, entry 3) and with the increase of the temperature
to 60 °C, the yield further improved to 85% with reduced
reaction time (Table 1, entry 4). Interestingly, when the
reaction was performed in methanol without catalytic acetic
acid at 60 °C the yield was slightly better (Table 1, entry 5).
Furthermore the reaction was also performed in ethanol and
dichloromethane, however the yields decreased compared
to methanol (Table 1, entries 6–7).
Results and discussion
Chemistry
The chemical strategy for the synthesis of the intermediate
amine 6 started with the condensation of 5-nitro-2-methyl
aniline (1) with cynamide to furnish the guanidine deriva-
tive (2), followed by reaction with pyridine based chalcone
(
4) in the presence of inorganic base to afford pyrimidine
derivative (5). The pyridine chalcone (4) was obtained from
-acetyl pyridine (3). The catalytic reduction of pyrimidine
3
derivative 5 with hydrogen on Pd/carbon in ethyl acetate
afforded the desired intermediate 6 (Scheme 1) (Zimmer-
mann 1993, 1996; Zimmermann et al. 1996).
Upon synthesizing 6, we proceeded towards the synth-
esis of the target compounds through imine formation. In

Medicinal Chemistry Research
Scheme 2 Reaction between
intermediate amine (6) with
isatin and aldehyde
Table 2 Synthesis of Imatinib based analogues
Moreover to further support the characterization of these
new conjugates, X-ray crystallography was performed with
compound 7a, which clearly identified the presence of
pyridine, pyrimidine and oxindole rings as well as the imine
bond with precision (Fig. 2).
Entry
R
Time (h)
Yield (%)
7
7
7
7
7
7
7
7
7
7
7
7
7
7
7
a
b
c
d
e
f
H
5
80
85
75
69
85
80
70
74
82
84
70
74
72
85
75
Cl
4.5
4
F
Br
5.5
6
Biological evaluation
I
Me
MeO
H
7
Inhibition of cell proliferation
g
h
i
7.5
6
To assess the potency of the novel conjugates of imitanib
(7a–7o), we evaluated their effects on cell proliferation
(Hamaï et al. 2006) as well as enzymatic activity with
imatinib drug as a reference. The decrease in cell viability of
K-562 cells treated with the compounds 7a–7o for 48 h
assessed with the in vitro SRB assay (Kasinski et al. 2015;
Skehan et al. 1990; Vichai and Kirtikara 2006) revealed
their promising anti-proliferative activity. All the deriva-
tives of the series (7a–7o) exhibited more potent cytotoxi-
city than imatinib against K-562 cell line in the range of
0.65–1.1 µM (Fig. 3). Compound 7l with indole ring
exhibited the best activity in the series with an IC50 value of
0.65 µM and almost two folds more potency than imitanib.
We believe introduction of XXX groups and specifically
XYY groups in ZZZ molecules enhanced their binding to
the kinase in focus and thus inhibiting cell proliferation.
Hence, to test the Abl kinase activity we consequently
performed in vitro kinase assays (Mow et al. 2002).
F
4.5
5
j
Cl
k
l
CN
NO2
MeO
CF3
F
5
4
m
n
o
6
6
6
Hence, we performed the condensation reaction of
intermediate 6 with different isatins and aldehydes under the
optimized reaction conditions with methanol as solvent at
6
0 °C to provide the desired conjugates 7a–7o as outlined in
Scheme 2. Equimolar mixture of compound 6 and different
isatins or aldehydes taken in methanol (5 ml) stirred at 60 °
C temperatures for 4–8 h furnished the desired compounds
in good to excellent yield (Table 2). Since this kind of
condensation reactions moves in the forward direction with
progressive removal of water, we carried out the reaction
with Dean and Stark apparatus with different water
immiscible solvents, however no remarkable changes were
observed in the yields and time.
Enzymatic kinase activity
In this study, all the compounds (7a–7o), were tested by
in vitro assay against the recombinant ABL kinase enzyme
(Ding et al. 2008; Ouellette et al. 2016). Most of them
displayed good anti-enzymatic activity with IC50 value
arranging from 30 to 71 µM, however under our experi-
mental conditions, imatinib drug showed a nine fold better
IC50 value. This indicates that there may be other molecular
targets where these compounds could bound and provides
better cytotoxicity values than imatinib. Further, com-
pounds 7a, 7h, 7n, and 7o showed no significant inhibitory
activity toward ABL (IC > 100 µM). Amongst the identi-
The structures of the final compounds were confirmed
1
13
based on their H, C NMR and HRMS data. In case of
1
compound 7a and 7b, we observed three singlet peaks in H
NMR spectra for isatinyl, pyridiny,l and secondary amine
protons at chemical shifts (δ) values of 10.85, 9.13, 8.64
and 11.07, 9.18, 8.93 respectively as well as in C NMR
1
3
characteristic peaks for compound 7a and 7b at δ 164.25,
1
63.72 for carbons directly attached to the three nitrogen, at
δ 161.89, 162.00 for carbons attached to imine bond, at δ
61.17, 161.23 for carbons attached to nitrogen of pyridine
ring and at δ 155.16, 154.03 for amide carbon respectively.
5
0
1
fied human kinome of 518 protein kinases many have been
identified to bind to same inhibitors thus ensuing

Medicinal Chemistry Research
Fig. 2 A view of 7a, showing
the atom-labeling scheme.
Displacement ellipsoids are
drawn at the 30% probability
level and H atoms are
represented by circles of
arbitrary radii
Fig. 3 In vitro cytotoxicity
against K562 cell line
polypharmacology (Manning et al. 2002). It has already
been known that kinase inhibitors show less promiscuity
towards their targets owing to their binding of homologous
ATP-binding sites in multiple freely available cellular
kinases. However their promiscuity is to be seen as ratio-
nalized due to their ability to bind to the same region of
several known protein kinases (Hu et al. 2017).
Structure activity relationship (SAR) can be explained
on the basis of values obtained from the cytotoxicity and
anti-enzymatic profile of compounds (7a–7o), (Fig. 3 and
Table 3). Interestingly, compounds having oxindole
(7a–7g) were found most active amongst the series; how-
ever there was no remarkable distinction in activity between
electron donating and withdrawing groups containing

Medicinal Chemistry Research
Table 3 In vitro bcr-abl enzymatic activity
Table 4 Drug-likeness properties of the top six active indole based
scaffolds
Compounds
IC50 (µM)
Compound TPSA
score
miLogP H-bond
acceptors
H-bond
donors
7
7
7
7
7
7
7
7
7
7
7
7
7
7
7
a
b
c
d
e
f
>100
60 ± 0.58
62 ± 0.64
71 ± 0.66
55 ± 0.42
40 ± 0.51
36 ± 0.38
>100
7
7
7
7
7
7
f
95.93
105.16
78.86
5.03
4.64
5.64
4.72
4.92
5.02
7
8
6
7
9
7
2
2
2
2
2
1
g
j
k
l
102.65
124.68
88.09
g
h
i
m
55 ± 0.49
37 ± 0.28
40 ± 0.36
30 ± 0.64
52 ± 0.32
>100
j
demonstrated vast therapeutic potential. The extent of
pharmacological activities ascribed to oxindole derivatives,
include anti-cancer, anti-viral anti-leishmanial, antibacterial,
antidiabetic, antioxidant, AChE inhibitory, adrenergic
receptor agonistic, analgesic, spermicidal, vasopressin/pro-
gesterone antagonistic, neuroprotection, and NMDA
blocker activities (Kaur et al. 2016). Oxindole derivatives
have gained respect in the field due to the principal com-
pound Sunitinib, that has been made for inhibition of
VEGFR2 specifically as an anti-angiogenesis compound
k
l
m
n
o
>100
Imatinib
3.7 ± 0.21
(Kang et al. 2016) that has been clinically approved for
renal cell carcinoma. Oxindole derivatives have been shown
to bind to ATP-binding region of the catalytic domain of
kinases.
7g
7j
7f
Druglikeness, toxicity and pharmacokinetic toxicity
and pharmacokinetics analysis
7
k
7l
7m
The pharmacokinetics and drug-likeness properties of the
hit compounds were studied using Mol-inspiration and
Swiss ADME online server by submitting SMILES. The
compounds were filtered out based on Water Partition
Coefficient (miLogP) values to be <5 which predict lower
toxicity, possible oral administration and non-specific
binding. The TPSA (Topological Polar Surface Area)
Fig. 4 Topological polar surface area (TPSA) of the top active syn-
thesized molecules
compounds. Nevertheless, methyl group slightly decreases
the activity with IC50 value 1.01 µM in compound 7f. In
case of indolyl compounds (7h–7m), electron rich indoles
have shown good activity however, halogenated and elec-
tron deficient indoles decreases the activity with an excep-
tion of nitro compound 7l which has shown excellent
activity with IC50 value 0.65 µM. Furthermore, aromatic
compounds (7n–7o), were found to have relatively similar
cytotoxicity as that of oxindole analogues (7a–7g) (Fig. 4).
In case of enzymatic activity, the oxindole analogues
0
score <140 A (da Silva et al. 2015) indicating a high
possibility of absorption (Table 4). In addition, the com-
pounds were also analyzed for bioavailability property
using Swiss ADME (Daina et al. 2017a, b) (Figs 4 and 5).
The pharmacokinetic and physicochemical properties of
all the synthesized compounds were studied using Swiss
ADME server. Among all the tested, five molecules were
selected as hit molecules by passing the filtering criteria of
(
7a–7g) having electron donating groups namely methyl
0
and methoxy have shown better activity than electron
withdrawing groups. However, in case of indole based
derivatives (7h–7m), electron deficient groups were found
to be better, particularly compound 7l with IC50 value 30
µM. None of the aromatic compounds (7n–7o) were found
to have anti-enzymatic activity.
miLogp < 5 and TPSA < 140 A . BOILED-Egg (Brain Or
IntestinaL EstimateD permeation method) is a perceptive
model to predict the passive gastrointestinal absorption
(HIA) and Blood Brain Barrier (BBB) permeation of small
molecules (Daina et al. 2017a, b). The BOILED-Egg ana-
lysis showed that most of the molecules (11 compounds) are
highly absorbable in the gastrointestinal tract. The active
efflux mechanism, P-gp as substrate has involved in the
Traditionally, oxindole extracted from the cat claw’s
plant Uncaria tomentosa in Amazon South America, have

Medicinal Chemistry Research
We confer that 7l may look as potential scope for lead
development.
Conclusion
Synthesis, characterization and in vitro evaluation of indole/
isatin conjugated phenyl-amino-pyrimidine derivatives as
potential BCR-ABL inhibitors have been demonstrated.
Among the series, all derivatives (7a–7o) were found to be
more cytotoxic than standard Imatinib against K-562 cell
line. Notably, compound 7l was the most active in the series
with almost two folds more potency than imitanib (IC50
0.65 μM). Furthermore, in vitro enzymatic studies with
recombinant ABL kinase enzyme exhibited promising
inhibition in the range of 30–71 µM for most of these novel
conjugates. In addition, modelling and other computational
studies have been carried out to draw insight into the BCR-
ABL protein interactions with the target molecules and drug
like properties of the conjugates, respectively. A detailed
study in understanding pattern of this biological activity
outcome would be worthwhile research investigation.
Fig. 5 Brain/intestinal permeation estimation
permeation of molecules (indicated as blue dots) to the
gastrointestinal lumen (Fig. 5).
Molecular docking studies
The molecular docking studies of 7h–7m designed inhibi-
tors demonstrated binding into the pocket of BCR-ABL
enzyme and forming one H-bond with the hinge region
Met-318 of Abl kinase catalytic domain, proven to be
essential for the inhibition of kinase activity (Parcha et al.
Experimental section
Chemistry
2
017; Banavath et al. 2014). Critical analysis of the ligand
All reagents, starting materials, and solvents were pur-
chased from Aldrich (Sigma-Aldrich, St. Louis, MO, USA)
or Alfa Aesar (Johnson Matthey Company, Ward Hill, MA,
USA) and 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 using an iodine indicator. Column
chromatography was performed with Merck 60–120 mesh
interaction pattern revealed that nitrogen of pyridyl ring
system involved in a H-bond with Met-318. The –NH linker
showed a H-bond with Tyr 315 and pyramidal ring exhib-
ited strong П-П stacking with Thr 253 as shown in sup-
plementary data with all synthesized molecules. Further
formation of salt bridge between Glu286, Asp 381 amino
acids and nitro substitution on indole moiety of compound
1
13
7
l contributed for BCR-ABL inhibition and absent in other
silica gel. H and C NMR spectra were recorded with 75,
derivatives. Interaction with Glu286, Asp 381 is an essential
interaction to show Bcr-Abl inhibition as suggested and
evidenced by Parcha et al. (2017) and also observed with
co-crystal ligand, nilotinib (Sabitha 2012) (Fig. 6).
From in vitro enzymatic and in silico studies it is evi-
denced that only compound 7l making interaction with
Glu286 and Asp 381 inhibited the BCR-ABL with an IC50
of 30 µM and arrested the K-562 cell growth with an IC50 of
100, 300, 400, and 500 MHz spectrometer in CDCl and
3
DMSO-d solutions. Chemical shifts (δ) are expressed in
6
ppm relative to the internal standard TMS and multiplicities
of NMR signals are represented as singlet (s), broad singlet
(bs), doublet (d), triplet (t), double doublet (dd), triplet of
doublet (td), and multiplets (m). High-resolution mass
spectra (ESI-HRMS) were obtained by using ESI-QTOF
mass spectrometer. Melting points were determined on an
electro-thermal melting point apparatus and are uncorrected.
0
.64 ± 0.09 demonstrating the potential lead like properties
of 7l to treat CML (Fig. 6).
It was observed that nilotinib formed hydrogen bonds
with Asp-381 and Glu-296 where as the compound 7l did
not formed these hydrogen bonds and this will be making
the compound more drug like as breaking the H-bonds
needs more energy making it physiologically non favorable
for the active site. Yet, the nitro group of the indole moiety
in 7l forms stable salt bridges with Asp-381 and Glu-296.
General procedure for the synthesis of compounds (7a–7o)
1
6-methyl-N -(4-(pyrdin-3-yl)pyrimidin-2-yl) benzene-1,3-
diamine (compound 6) (0.5 mmol) and 5-substituted, indo-
line-2,3-dione(isatins) or indole and aromatic aldehydes
(0.5 mmol) were taken in methanol (5 ml) and stirred at
60 °C temperature for 4–8 h. The reaction was monitored by

Medicinal Chemistry Research
Fig. 6 Binding pose of 7l and nilotinib into the active site pocket of BCR-ABL enzyme
TLC using methanol and chloroform (2%) as a solvent
system. After completion of reaction this mixture was
evaporated by vacuum distillation and the residue was
extracted with ethyl acetate (25 mL × 3). The organic solu-
tion was dried over anhydrous Na SO and evaporated the
145.92, 139.08, 134.33, 133.86, 132.45, 131.50, 128.89,
125.95, 125.65, 123.71, 117.03, 113.78, 113.57, 113.12,
108.11, 18.11; HRMS calculated for C H ON Cl
2
4
18
6
+
[M + H] 441.1225; found: 441.1225; C H ON Cl [M +
2
4
18
6
+
2] 443.1196; found: 443.1236;
2
4
solvent to offered crude product. This was further purified
by column chromatography using 0.1:10 methanol/chloro-
form as eluent to obtain the pure compound in good yield.
(Z)-5-Fluoro-3-((4-methyl-3-((4-pyrdin-3-yl) pyrimidin-2yl)
amino) phenyl) imino) indolin-2-one (7c) White solid
1
(
75%); R : 0.28 (5% MeOH/CHCl ); M.P.: 243–245 °C, H
f
3
(
Z)-3-((4-Methyl-3-((4-pyrdin-3-yl) pyrimidin-2-yl) amino-
NMR (300 MHz, CDCl + DMSO-d ) δ 10.78 (s, 1H), 9.08
3
6
phenyl) imino) indolin-2-one (7a) White solid (80%); R :
(s, 1H), 8.54 (d, J = 3.9 Hz, 1H), 8.36 (d, J = 5.2 Hz, 1H),
8.17 (d, J = 8.0 Hz, 1H), 7.53 (s, 1H), 7.26–7.07 (m, 5H),
f
1
0
.35 (5%, MeOH/CHCl ); M.P.: 254-255 °C; H NMR
3
(
300 MHz, CDCl + DMSO-d ) δ 10.85 (s, 1H), 9.13 (s,
6.94–6.79 (m, 1H), 6.66–6.63 (m, 2H), 2.33 (s, 3H).; 13
C
3
6
1
4
7
1
H), 8.64 (s, 1H), 8.57 (d, J = 3.9Hz, 1H), 8.40 (d, J =
.9Hz, 1H), 8.20 (d, J = 7.7 Hz, 1H), 7.37 (s, 1H), 7.29–
.16 (m, 4H), 6.82 (dd, J = 12.5, 7.8 Hz, 2H), 6.62 (dd, J =
6.5, 8.2 Hz, 2H), 2.32 (s, 3H).; 1 C NMR (75 MHz,
NMR (75 MHz, CDCl + DMSO-d ) δ 164.48, 162.68 (d,
J1 = 270.7 Hz, 1C), 158.97, 155.88, 154.41, 151.10,
148.16, 147.73, 143.22, 138.58, 134.30, 132.51, 131.29,
3
6
3
127.27, 123.48, 120.48 (d, J = 26.5 Hz, 1C), 116.22
2
CDCl + DMSO-d ) δ 164.25, 161.89, 161.17, 159.25,
(d, J = 7.8 Hz, 1C), 113.82, 113.35, (d, J = 26.7 Hz, 1C),
3
6
3
2
1
1
1
55.16, 151.25, 148.51, 148.23, 147.16, 138.90, 134.27,
32.44, 131.33, 127.99, 126.30, 123.60, 121.86, 115.98,
112.37, 108.06, 17.89; HRMS (ESI) calculated for
+
C H ON F [M + H] 425.1520; found: 425.1512.
2
4
18
6
13.63, 113.39, 111.57, 107.90, 18.00. HRMS (ESI) cal-
+
culated for C H ON [M + H] 407.1614; found:
(Z)-5-Bromo-3-((4-methyl-3-((4-pyrdin-3-yl) pyrimidin-2-yl)
2
4
19
6
4
07.1613.
amino) phenyl) imino) indolin-2-one (7d) White solid
1
(
69%); R : 0.30 (5% MeOH/CHCl ); M.P.: 194-196 °C; H
f
3
(
Z)-5-Chloro-3-((4-methyl-3-((4-pyrdin-3-yl) pyrimidin-2-yl)
NMR (300 MHz, CDCl + DMSO) δ 10.83 (s, 1H), 9.16 (s,
3
amino) phenyl) imino) indolin-2-one (7b) White solid
1H), 8.64 (d, J = 4.6 Hz, 1H), 8.48 (t, J = 4.4 Hz, 1H), 8.24
(d, J = 8.1 Hz, 1H), 7.82 (s, 1H), 7.77 (s, 1H), 7.59 (s, 1H),
7.34–7.28 (m, 2H), 7.19 (d, J = 5.1 Hz, 1H), 7.15 (s, 1H),
6.80 (t, J = 8.6 Hz, 1H), 6.74 (d, J = 7.7 Hz, 1H), 2.44 (s,
1
(
85%); R : 0.25 (5% MeOH/CHCl ); M.P.: 252-253 °C; H
f 3
NMR (300 MHz, CDCl + DMSO-d ) δ 11.07 (s, 1H), 9.18
3
6
(
5
s, 1H), 8.93 (s, 1H), 8.61 (d, J = 3.8Hz, 1H), 8.44 (d, J =
.1Hz, 1H), 8.29 (d, J = 7.9 Hz, 1H), 7.32 (m, 5H), 6.88 (d,
J = 8.4 Hz, 1H), 6.77 (s, 1H), 6.70 (d, J = 7.4 Hz, 1H), 2.34
1
3
3H); C NMR (75 MHz, CDCl + DMSO) δ 163.2, 161.7,
3
160.9, 159.44, 153.7, 151.31, 148.1, 147.7, 145.9, 138.8,
136.5, 134.2, 132.3, 132.1, 131.3, 130.4, 128.8, 123.6,
119.5, 117.3, 113.5, 111.1, 107.9, 17.8; HRMS (ESI)
1
3
(
s, 3H); C NMR (75 MHz, CDCl + DMSO-d ) δ 163.72,
3
6
1
62.00, 161.23, 159.42, 154.03, 151.42, 148.35, 147.90,

Medicinal Chemistry Research
+
calculated for C H ON Br [M + H] 485.0720; found:
6.4 Hz, 2H), 8.23 (s, 1H), 8.03 (d, J = 7.0 Hz, 1H), 7.65–7.50
(m, 2H), 7.44 (s, 2H), 7.29–7.13 (m, 3H), 6.94 (d, J = 7.3 Hz,
2
4
18
6
+
4
85.0748; C H ON Br [M + 2]
485.0700; found:
2
4
18
6
1
3
4
85.0727.
1H), 2.21 (s, 3H); C NMR (100 MHz, DMSO-d ) δ 185.51,
6
162.01, 161.18, 160.03, 151.44, 148.18, 139.20, 138.96,
(
Z)-5-Iodo-3-((4-methyl-3-((4-pyrdin-3-yl)
pyrimidin-2-yl)
137.52, 135.58, 131.70, 130.81, 130.66, 124.67, 123.94,
122.61, 121.28, 118.31, 112.92, 108.74, 18.18; HRMS (ESI)
calculated for C H N [M + H]+ 405.1822; found:
amino) phenyl) imino) indolin-2-one (7e) White solid
(
1
85%); R : 0.40 (5% MeOH/CHCl ); M.P.: 126-128 °C; H
f 3
25 21 6
NMR (300 MHz, CDCl + DMSO-d ) δ 10.91 (s, 1H), 9.09
405.1822.
3
6
(
8
1
s, 1H), 8.54 (d, J = 3.4 Hz, 1H), 8.40 (d, J = 5.2 Hz, 1H),
1
3
.17 (d, J = 8.0 Hz, 1H), 7.80 (d, J = 2.8 Hz, 1H), 7.57 (s,
H), 7.45 (d, J = 8.2 Hz, 1H), 7.24 (d, J = 8.0 Hz, 1H), 7.17
(E)-N ((5-Fluoro-1H-indol-3-yl)methylene)-4-methyl-N -(4-
(pyridine-3-yl) pyrimidin-2-yl) benzene-1,3-diamine (7i)
White solid (82%); R : 0.24 (5% MeOH/CHCl ); M.P.:
13
(
d, J = 5.5 Hz, 1H), 2.35 (s, 3H); C NMR (75 MHz,
f
3
1
CDCl + DMSO-d ) δ 163.74, 162.24, 162.06, 160.86,
146–148 °C; H NMR (500 MHz, DMSO-d ) δ 12.38 (s,
3
6
6
1
1
1
1
1
5
59.24, 159.12, 153.56, 151.12, 148.23, 147.74, 146.58,
46.28, 145.86, 138.66, 134.51, 133.13, 132.61, 131.27,
30.77, 127.34, 123.75, 123.57, 117.91, 115.00, 113.85,
13.64, 112.26, 111.16, 110.11, 108.25, 107.54, 84.16,
1H), 9.98 (s, 1H), 9.41 (s, 1H), 9.20 (s, 1H), 8.84 (dd, J =
14.3, 4.1 Hz, 1H), 8.72 (d, J = 7.9 Hz, 1H), 8.65 (t, J = 4.6
Hz, 1H), 8.41 (d, J = 3.1 Hz, 1H), 7.81 (dd, J = 14.2, 7.3
Hz, 3H), 7.60 (dd, J = 9.7, 4.9 Hz, 2H), 7.41 (d, J = 8.1 Hz,
1H), 7.19 (dd, J = 9.2, 2.5 Hz, 1H), 7.16–7.12 (m, 1H), 2.35
+
8.01; HRMS (ESI) calculated for C H ON I [M + H]
24
18
6
1
3
33.0586; found: 533.0575.
(s, 3H); C NMR (100 MHz, DMSO-d ) δ 185.54, 161.37,
6
1
61.29, 161.10, 160.23, 159.16 (d, J = 234.9Hz, 1C),
1
(
Z)-5-Methyl-3-((4-methyl-3-((4-pyrdin-3-yl) pyrimidin-2-yl)
149.61, 146.55, 140.10, 139.32, 139.16, 137.58, 137.40,
134.07, 133.43, 131.78, 131.68, 130.03, 125.47, 119.04,
114.27 (d, J = 9.7 Hz, 1C), 112.07 (d, J = 25.9Hz, 1C),
amino) phenyl) imino) indolin-2-one (7f) White solid
(
1
80%); R : 0.20 (5% MeOH/CHCl ); M.P.: 134-136 °C; H
f 3
3
2
NMR (300 MHz, CDCl + DMSO-d ) δ 10.35 (s, 1H), 9.05
108.94, 106.13 (d, J = 24.5 Hz, 1C), 18.21; HRMS (ESI)
3
6
2
(
s, 1H), 8.54 (d, J = 3.7 Hz, 1H), 8.36 (d, J = 5.1 Hz, 1H),
calculated for C H FN [M+H]+ 423.1732; found:
2
5
21
6
8
2
6
.12 (d, J = 7.9 Hz, 1H), 7.71 (s, 1H), 7.53 (s, 1H), 7.44 (s,
H), 7.09 (d, J = 5.1 Hz, 1H), 6.95 (d, J = 7.9 Hz, 1H),
.76–6.56 (m, 3H), 2.34 (s, 3H), 1.87 (s, 3H); 1 C NMR
423.1728.
3
(E)-3-(((4-Methyl-3-((4-(pyridin-3-yl)pyrimidin-2-yl)amino)
phenyl)imino)methyl)-1H-indole-5-carbonitrile (7k) White
solid (70 %); R : 0.22 (5% MeOH/CHCl ); M.P.: 231–233 °C;
(
75 MHz, CDCl + DMSO-d ) δ 164.72, 162.15, 160.62,
3
6
1
1
1
58.86, 154.97, 151.04, 148.41, 148.04, 144.49, 143.87,
38.11, 134.33, 130.96, 126.78, 125.76, 123.33, 113.38,
f
3
1
H NMR (400 MHz, DMSO-d ) δ 12.78 (s, 1H), 9.98 (s, 1H),
6
11.73, 111.08, 107.99, 20.61, 17.71; HRMS (ESI) calcu-
9.47 (s, 1H), 9.24 (s, 1H), 8.98 (d, J = 7.9 Hz, 1H), 8.93 (s,
1H), 8.63 (d, J = 4.9 Hz, 1H), 8.50 (d, J = 2.9 Hz, 1H), 8.44
+
lated for C H ON [M + H] 421.1771; found: 421.1763.
2
5
21
6
(s, 1H), 7.99 (dd, J = 15.0, 9.4 Hz, 1H), 7.79 (d, J = 1.4 Hz,
(
Z)-5-Methoxy-3-((4-methyl-3-((4-pyrdin-3-yl)pyrimidin-2-yl)
1H), 7.71 (d, J = 8.5 Hz, 1H), 7.66–7.58 (m, 2H), 7.35 (d, J
amino)phenyl)imino)indolin-2-one (7g) White solid (70%);
= 8.1 Hz, 1H), 7.11 (dd, J = 8.0, 1.7 Hz, 1H), 2.28 (s, 3H);
1
13
R : 0.22 (5% MeOH/CHCl ); M.P.: 132–134 °C; H NMR
C NMR (100 MHz, DMSO-d ) δ 185.97, 161.01, 160.51,
f
3
6
(
300 MHz, DMSO-d ) δ 10.41 (s, 1H), 9.05 (s, 1H), 8.53 (d,
160.07, 146.61, 143.91, 140.77, 139.33, 138.97, 134.60,
131.80, 129.82, 126.89, 126.77, 126.15, 124.41, 120.42,
119.28, 118.42, 114.50, 109.12, 104.82, 18.20; HRMS (ESI)
6
J = 3.7 Hz, 1H), 8.36 (d, J = 5.1 Hz, 1H), 8.12 (d, J = 7.9 Hz,
H), 7.81 (s, 1H), 7.61 (d, J = 1.8 Hz, 1H), 7.54 (d, J = 3.5
Hz, 1H), 7.21 (d, J = 8.1 Hz, 1H), 7.10 (d, J = 5.1 Hz, 1H),
.72 (s, 2H), 6.62 (dd, J = 7.9, 1.8 Hz, 1H), 6.37 (s, 1H), 3.32
s, 3H), 2.32 (s, 3H); 13C NMR (75 MHz, CDCl + DMSO-
1
+
calculated for C H N [M+H] 430.1782; found: 430.1774.
25 21 6
6
(
1
1
(E)-4-methyl-N -((5-nitro-1H-indol-3-yl)methylene)-N -(4-
3
d6): δ 164.0, 161.4, 158.2, 153.7, 150.3, 147.4, 147.3, 144.1,
(pyridine-3-yl)pyrimidin-2-yl)benzene-1,3-diamine
(7l)
139.8, 137.6, 133.7, 133.6, 130.4, 130.0, 124.5, 122.8, 118.9,
White solid (74%); R : 0.20 (5% MeOH/CHCl ); M.P.:
f
3
1
1
12.8, 111.3, 110.2, 107.3, 54.5, 17.0; HRMS (ESI) calcu-
244–245 °C; H NMR (300 MHz, CDCl + DMSO-d ) δ
3
6
+
lated for C H O N [M + H] 437.1720; found: 437.1714.
10.04 (s, 1H), 9.31 (d, J = 2.0 Hz, 2H), 9.02 (s, 1H), 8.85 (s,
H), 8.71 (s, 2H), 8.55 (d, J = 5.1 Hz, 1H), 8.45 (dd, J =
2
5
21
2 6
1
1
3
(
3
E)-N ((1H-indol-3-yl) methylene)-4-methyl-N -(4-(pyridine-
-yl) pyrimidin-2-yl) benzene-1,3-diamine (7h) White
solid (74%); R : 0.33 (5% MeOH/CHCl ); M.P.: 126–128 °C;
17.2, 7.1 Hz, 2H), 8.26 (s, 1H), 8.14 (dd, J = 8.9, 2.2 Hz,
1H), 7.69 (d, J = 9.0 Hz, 1H), 7.59 (t, J = 3.1 Hz, 1H), 7.54
(dd, J = 7.5, 4.7 Hz, 1H), 7.46 (d, J = 5.1 Hz, 1H), 7.38 (d,
J = 5.1 Hz, 1H), 7.30 (d, J = 8.1 Hz, 1H), 7.06 (dd, J = 8.0,
1.8 Hz, 1H), 6.89 (d, J = 8.1 Hz, 1H), 6.81 (d, J = 1.9 Hz,
f
3
1
H NMR (300 MHz, DMSO-d ) δ 12.12 (s, 1H), 9.87 (s, 1H),
6
9.23 (s, 1H), 9.00 (s, 1H), 8.66 (s, 1H), 8.47 (dd, J = 15.9,

Medicinal Chemistry Research
1
3
1
1
1
H), 6.36 (dd, J = 8.0, 2.1 Hz, 1H), 4.95 (s, 1 H), 2.30 (s,
line was seeded in media containing 10% FBS and based
on the cells replicative capacity, plating was performed.
Prior to treatment with experimental drugs, plates were
1
3
H); C NMR (75 MHz, CDCl + DMSO-d ) δ 164.0,
3
6
61.7, 160.4, 158.5, 153.9, 150.6, 147.7, 147.3, 142.7,
38.1, 133.8, 132.0, 130.8, 123.0, 120.2, 119.8, 115.8,
maintained at 100% relative humidity, 5% CO , 95% air
2
13.3, 113.0, 112.7, 111.9, 107.6, 17.4; HRMS (ESI) cal-
and temperature of 37 °C. To each well, 198 μL of med-
ium containing cells and 2 μL of test compounds were
added. The assay was performed in triplicates for each
sample and five different concentrations (0.01, 0.1, 1, 10,
and 100 μM) were taken into consideration. After incu-
bation for 24 h, 100 μL of ice-cold 10% TCA was added
and incubated at 40 °C for 60 min. Post-incubation plates
were washed thrice with water and dried at room
temperature.
+
culated for C H N [M + H] 450.1682; found: 450.1673.
2
5
21
6
3
1
(
E)-4-methyl-N -(4-pyrdin-3-yl)pyrimidin-2-yl)-N -((4-tri-
fluoromethylbenzylidene) benzene-1,3-diamine (7n)
White solid (85%); R : 0.27 (5% MeOH/CHCl ); M.P.:
1
9
8
f
3
1
22–124 °C; H NMR (300 MHz, CDCl + DMSO-d ): δ
3
6
.22 (d, J = 1.5 Hz, 1H), 8.63 (dd, J = 4.8, 1.4 Hz, 1H),
.55 (s, 1H), 8.44 (d, J = 5.2 Hz, 1H), 8.27 (dd, J = 6.2, 1.9
Hz, 1H), 7.98 (d, J = 8.5 Hz, 3H), 7.65 (d, J = 8.1 Hz, 2H),
.50 (s, 1H), 7.34 (dd, J = 7.7, 4.9 Hz, 1H), 7.19 (d, J = 8.1
Hz, 1H), 7.13 (d, J = 5.2 Hz, 1H), 6.91 (dd, J = 8.0, 2.0 Hz,
H), 2.31 (s, 3H). 13C NMR (75 MHz, CDCl + DMSO-
To each well, 100 μL of SRB (sulforhodamine B)
solution was added and incubated at room temperature
for 30 min. Plates were washed with 1% acetic acid
thrice to remove unbound dye and left for drying over-
night. A volume of 200 μl of 10 mM Tris base solution
was added to each well to solubilize the protein bound
dye and measurements were taken at 510 nm with aid of
Varioscan Flash multimode plate reader. Based on
the percentage growth in controls, the inhibitory con-
centrations were determined and graphs were plotted
between percentage of viable cells and concentration of
compounds. IC50 values were calculated and reported as
result of mean ± SD for all triplicate independent
experiments.
7
1
3
d6): δ 162.10, 160.83, 159.12, 158.99, 157.57, 151.24,
1
1
1
1
1
49.19, 148.31, 139.46, 138.31, 137.87, 134.31, 132.54, δ
31.88 (q, J = 32.1 Hz, 1C), 131.24, 130.93, 130.72,
29.90, 128.92, 128.45, 125.95 (q, J = 3.6 Hz, 1C), 125.50,
25.46, 123.88 (q, J = 272.2 Hz, 1C), 123.55, 116.84,
15.07, 110.88, 109.61, 107.97, 107.50, 17.88; HRMS
+
(
ESI) calculated for C H N F [M + H] 434.1582;
24
19
5 3
found: 434.1587.
1
3
(
E)-N -(4-fluorobenzylidene)-4-methyl-N -(4-pyrdin-3-yl)pyr-
imidin-2-yl)benzene-1,3-diamine (7o) White solid (75%);
1
R : 0.32 (5% MeOH/CHCl ); M.P.: 138–140 °C; H NMR
BCR-ABL kinase inhibitory assay
f
3
(
300 MHz, CDCl + DMSO-d ): δ 9.20 (s, 1H), 8.62 (s,
3
6
1
7
2
1
1
1
1
1
3
3
H), 8.45 (s, 1H), 8.28 (d, J = 5.5 Hz, 1H), 7.97 (s, 2H),
The kinase inhibitory effect of the test compounds was
assessed by Z-lyte kinase assay kit (details) according to
manufacturer’s instructions which is based on the differ-
ential sensitivity of phosphorylated and non-
phosphorylated peptides to proteolytic cleavage. Test
compounds with 5 mM concentrations was diluted to
1000 μM with DMSO and transferred to the dose plate.
Further, each compound was diluted to 10-fold con-
centration with reaction buffer to obtain a 10X final
concentration. Later, the compounds (1 µL/well) were
transferred to assay plate with a concentration ranging
from 100 to 0.006 μM for the BCR-ABL activity. Imatinib
was run in parallel as a positive control. Reaction mixture
with ATP (0% inhibition), without ATP (100% inhibition)
and without ATP and kinase peptide mixture (100%
phosphorylation) are used as controls in the experiments.
The reaction mixtures were mixed and incubated 1 h at
room temperature followed by development reaction. All
the plates were incubated at room temperature for 1 h and
the fluorescence signal was measured at an excitation and
emission wavelengths of 400 and 445 nm, respectively.
Each experiment was performed in triplicates and the
results are expressed as mean ± S.D.
.85 (s, 1H), 7.50 (m, 2H), 7.20–6.97 (m, 5H), 6.88 (s, 1H),
1
3
.30 (s, 3H); C NMR (75 MHz,CDCl + DMSO-d ): δ
3
6
67.48, δ 164.44 (d, J = 252.2 Hz), 162.35, 160.73,
1
59.06, 158.12, 151.24, 149.99, 148.31, 138.04, 134.52,
32.65, 132.26, 130.94, 130.68, 127.26, 127.02, 123.64,
16.73, 116.29 (d, J = 22.6 Hz), 115.93, 115.64, 115.37,
2
15.08, 114.27, 110.92, 109.12, 17.79; MS–ESI-MS: m/z
+
84 HRMS (ESI) calculated for C H N O [M + H]
25
21
6
84.1626; found: 384.1619.
Biology
Cell cultures, maintenance, and evaluation of anti-
proliferative activity
Cell line utilized during the experiments was procured
from American Type Culture Collection (ATCC, USA),
K562 cells were grown in RPMI1640 medium at 37 °C
containing 10% Fetal Bovine Serum (FBS). The cells
were collected after 24 h post treatment. To estimate the
cell growth in the presence of the compounds, SRB cell
proliferation assay was performed. In a 96 well plate, cell

Medicinal Chemistry Research
Molecular docking studies
CB2 1EZ, UK; fax: +44(0) 1223 336 033; email:
deposit@ccdc.cam.ac.uk].
All computational calculations were carried out on an Intel
Acknowledgements The author AR thanks CSIR and UGC, New
Delhi for the award of fellowships. The author RS thanks CSIR-
HRDG for the award of CSIR-SRAship (13(8906-A)/2017-pool).
(
R) Xenon(R) 2 Duo CPU E7600 @ 3.06 GHz processor
with memory of 2 GB RAM running with the LINUX
operating system. Software package used was Schrodinger
suite 2017 drug discovery suite. Docking (Friesner et al.
Compliance with ethical standards
2
006) studies were perfomed for dataset compounds on to
the active site of Bcr-Abl crystal structure (PDB ID: 3CS9)
using Schrodinger Suite 2017. Structures were sketched and
converted to 3D using Ligprep (Schrödinger 2014). Bcr-Abl
crystal structure was prepared using Protein Prep wizard,
Schrodinger Suite. Further grid was defined using center of
nilotinib co-crystal as reference. Top 10 docking poses for
each compound was generated and analyzed. Manual
inspection of hits was performed to understand the binding
pattern and docking scores were considered.
Conflict of interest The authors declare that they have no conflict of
interest.
Publisher’s note: Springer Nature remains neutral with regard to
jurisdictional claims in published maps and institutional affiliations.
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.5U (C) for methyl H or 1.2U (C) for other H atoms]. The
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−1 0, −0.995 −0.006 −0.989. The structure was refined
using the hklf 5 routine with all reflections.
Crystal data for 7a C H N O (M = 406.44 g/mol): tri-
2
4
18
6
clinic, space group P-1 (no. 2), a = 8.46500(10) Å, b =
1
1.00000(10) Å, c = 11.4730(3) Å, α = 70.5200(6)°, β =
3
8
0.6100(6)°, γ = 89.3400(7)°, V = 992.60(3) Å , Z = 2,
T = 294.15 K, μ(MoKα) = 0.088 mm-1, D = 1.360 g/
calc
3
cm , 11,518 reflections measured (4.468° ≤ 2Θ ≤ 50°),
1
1518 unique (Rint = ?, Rsigma = 0.0976) which were used
in all calculations. The final R was 0.0813 (I > 2σ(I)) and
1
wR was 0.2450 (all data). CCDC 1841348 contains sup-
2
osine kinase inhibitors in chronic myeloid leukemia:
a
plementary Crystallographic data for the structure. These
data can be obtained free of charge at www.ccdc.cam.ac.uk/
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