Synthesis, X-ray, Hirshfeld surface analysis, exploration of DNA binding, urease enzyme inhibition and anticancer activities of novel adamantane-naphthyl thiourea conjugate

Article abstract of DOI:10.1016/j.bioorg.2021.104707

1-(adamantane-1-carbonyl-3-(1-naphthyl)) thiourea (C₂₂H₂₄N₂OS (4), was synthesized by the reaction of freshly prepared adamantane-1-carbonyl chloride from corresponding acid (3) with ammonium thiocyanate in 1:1 M ratio in dry acetone to afford the adamantane-1-carbonyl isothiocyanate (2) in situ followed by treatment with 1-naphthyl amine (3). The structure was established by elemental analyses, FTIR, ¹H, ¹³C NMR and mass spectroscopy. The molecular and crystal structure were determined by single crystal X-ray analysis. It belongs to triclinic system P ? 1 space group with a = 6.7832(5) ?, b = 11.1810(8) ?, c = 13.6660(10) ?, α = 105.941(6)°, β = 103.730(6)°, γ = 104.562(6)°, Z = 2, V = 910.82(11) ?³. The naphthyl group is almost planar. In the crystal structure, intermolecular C[sbnd]H···O hydrogen bonds link the molecules into centrosymmetric dimers, enclosing R₂²(14) ring motifs, while the intramolecular N[sbnd]H···O hydrogen bonds enclose S(6) ring motifs, in which they may be effective in the stabilization of the structure. The Hirshfeld surface analysis of the crystal structure indicates that the most important contributions for the crystal packing are from H … H (59.3%), H … C/C … H (19.8%) and H … S/S … H (10.1%) interactions. Hydrogen bonding and van der Waals interactions are the dominant interactions in the crystal packing. DFT, molecular docking and urease inhibition studies revealed stability and electron withdrawing nature of 4 as compared to DNA base pairs and residues of urease. The DNA binding results from docking, UV– visible spectroscopy, and viscosity studies indicated significant binding of 4 with the DNA via intercalation and groove binding. Further investigation of the compound was done on hepatocellular carcinoma; Huh-7 cell line as well as normal human embryonic kidney; Hek-293 cell line. The compound showed significant cytotoxic activity against Huh-7 cells in comparison to normal Hek-293 cells indicating selective cytotoxicity towards cancer cells.

Full text of DOI:10.1016/j.bioorg.2021.104707

Bioorganic Chemistry 109 (2021) 104707
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Synthesis, X-ray, Hirshfeld surface analysis, exploration of DNA binding,
urease enzyme inhibition and anticancer activities of novel
adamantane-naphthyl thiourea conjugate
a
,
*
b
,
*
c
d
a
Nasima Arshad , Aamer Saeed , Fouzia Perveen , Rabail Ujan , Shahid I. Farooqi ,
b
b
e
f
f
Pervaiz Ali Channar , Ghulam Shabir , Hesham R. El-Seedi , Aneela Javed , Maham Yamin ,
Michael Bolte , Tuncer H o¨ kelek
g
h
a
Department of Chemistry, Allama Iqbal Open University, 44000 Islamabad, Pakistan
b
c
d
e
f
Department of Chemistry, Quaid-i-Azam University, 45320 Islamabad, Pakistan
Research Center for Modeling and Simulations, National University of Sciences and Technology (NUST), Islamabad, Pakistan
M. A. Kazi Institute of Chemistry, University of Sindh, Jamshoro 76080, Pakistan
Pharmacognosy Group, Department of Medicinal Chemistry, Biomedical Center (BMC), Uppsala University, Uppsala, Sweden
Healthcare Biotechnology, Atta-ur-Rehman School of Applied Biosciences, National University of Science and Technology (NUST), Islamabad, Pakistan
Institut für Anorganische Chemie, J.W.-Goethe-Universit a¨ t, Max-von-Laue-Str.7, D-60438 Frankfurt/Main, Germany
Hacettepe University, Department of Physics, 06800 Beytepe-Ankara, Turkey
g
h
A R T I C L E I N F O
A B S T R A C T
Keywords:
1-(adamantane-1-carbonyl-3-(1-naphthyl)) thiourea (C22H24N OS (4), was synthesized by the reaction of freshly
2
Adamantanyl thiourea
X-ray
prepared adamantane-1-carbonyl chloride from corresponding acid (3) with ammonium thiocyanate in 1:1 M
ratio in dry acetone to afford the adamantane-1-carbonyl isothiocyanate (2) in situ followed by treatment with 1-
naphthyl amine (3). The structure was established by elemental analyses, FTIR, 1H, 13C NMR and mass spec-
Hirshfeld surface analysis
DNA binding
troscopy. The molecular and crystal structure were determined by single crystal X-ray analysis. It belongs to
Urease inhibition
Anticancer potential
◦
triclinic system P ꢀ 1 space group with a = 6.7832(5) Å, b = 11.1810(8) Å, c = 13.6660(10) Å,
α
= 105.941(6) ,
◦
◦
3
β = 103.730(6) , γ = 104.562(6) , Z = 2, V = 910.82(11) Å . The naphthyl group is almost planar. In the crystal
structure, intermolecular C
H⋅⋅⋅O hydrogen bonds link the molecules into centrosymmetric dimers, enclosing
(14) ring motifs, while the intramolecular N
H⋅⋅⋅O hydrogen bonds enclose S(6) ring motifs, in which they
may be effective in the stabilization of the structure. The Hirshfeld surface analysis of the crystal structure in-
dicates that the most important contributions for the crystal packing are from H … H (59.3%), H … C/C …
19.8%) and H … S/S … H (10.1%) interactions. Hydrogen bonding and van der Waals interactions are the
–
2
2
R
–
H
(
dominant interactions in the crystal packing. DFT, molecular docking and urease inhibition studies revealed
stability and electron withdrawing nature of 4 as compared to DNA base pairs and residues of urease. The DNA
binding results from docking, UV– visible spectroscopy, and viscosity studies indicated significant binding of 4
with the DNA via intercalation and groove binding. Further investigation of the compound was done on hepa-
tocellular carcinoma; Huh-7 cell line as well as normal human embryonic kidney; Hek-293 cell line. The com-
pound showed significant cytotoxic activity against Huh-7 cells in comparison to normal Hek-293 cells indicating
selective cytotoxicity towards cancer cells.
1
. Introduction
heterocyclic compounds and exhibit a broad spectrum of bioactivities
′
[
1,2]. Thus, 1-acyl-3-(2 -aminophenyl) thioureas are anti-intestinal
There has been an unprecedented upsurge in the synthesis of various
,3-disubstituted thioureas in recent years. Thiourea derivatives are
nematode prodrugs [3]; the thiourea derivatives of 4-azatricyclo
[5.2.2. 02,6] undec-8-ene-3,5-dione [2] and pyridazine [4]; 1-aroyl-3-
arylthioureas [5], Jack bean urease inhibitors and 1-aroyl-3-
1
exceptionally versatile building blocks for the synthesis of a diversity of
*
Corresponding authors.
E-mail addresses: nasima.arshad@aiou.edu.pk (N. Arshad), asaeed@qau.edu.pk (A. Saeed).
https://doi.org/10.1016/j.bioorg.2021.104707
Received 28 September 2020; Received in revised form 10 December 2020; Accepted 27 January 2021
Available online 9 February 2021
0
045-2068/© 2021 Elsevier Inc. All rights reserved.

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
(
substituted-2-benzothiazolyl) thioureas exhibit potent antibacterial
work was the synthesis of new adamantyl bearing thiourea to combine
its valuable effect in a single structural entity. Besides, X-ray single
crystal, Hirshfeld surface analysis, DFT, urease inhibition, DNA binding
and anticancer activity studies have been carried out to explore the
marvelous characteristics of the synthesized compound.
activity [6]. N-Phenyl and N-benzoylthioureas are antimicrobial agents
[
6] and the fluorinated thioureas exhibit better antifungal than the
′
antibacterial activity [7]. N-phenyl-N -[4-(5-cyclohexyl amino-1,3,4-
thiadiazole-2-yl)phenyl]thioureas, [8], 1-(5-cyclobutyl-1,3-oxazol-2-
′
′
yl)-3(sub)phenyl/pyridyl thioureas [9] and N-pentofuranosyl-N -[p-
isoamyloxy)phenyl] thioureas are potent anti-tuberculosis therapeutic
agents toward multidrug-resistant Mycobacterium tuberculosis [10]. N-D-
2. Experimental
′
Aldopentofuranosyl-N -[p-(isoamyloxy) phenyl] thioureas, designed as
2.1. Materials and methods
structural analogues of thiocarlide, are even more potent than thio-
carlide itself [11]. 6-Thioureido-4-anilinoquinazolines are antimalarial
agents [12], thioureas derived from 4-aminobenzohydrazide hydra-
zones show antiviral, antitubercular and anticancer activities [13]. In
addition, N-4-substituted benzyl-N’-ter-butyl benzylthioureas show an-
Thin layer chromatography (TLC) was conducted on 0.25 mm silica
gel plates (60 F254, Merck). Visualization was made with ultraviolet
light. Reagents were obtained commercially and used as received. By
using Falcon protocol, double-strand (ds) DNA was extracted in the
laboratory from the calf thymus gland. In order to check the purity of
DNA, the DNA threads were dissolved in autoclave distilled water
overnight. Absorption spectra of DNA solution was monitored by UV–
visible spectrophotometer and absorbance ratio (A260/A280) was calcu-
lated up to 1.89 which ensures that the DNA solution is sufficiently pure
and has no protein content. The concentration of the stock DNA solution
algesics
activity
[14],
3,4-dimethoxy
phenylethyl-1,3,5-
triazinylthioureas exhibit anti-HIV activity [15]; polysubstituted
acylthioureas are a novel class of potent influenza virus inhibitors [16],
while still other thioureas are novel insecticides for control of the
dengue vector [17], N-(o-fluorophenoxyacetyl) thioureas possess her-
bicidal agents [18] and plant growth regulators [19].
ꢀ
1
ꢀ 1
Conversely the use of the adamantyl group in drug design is also
continuously escalating. Adamantyl-based compounds are used for the
treatment of neurological conditions, as antiviral agents and as agents
against type 2 diabetes. The value of the adamantyl group in drug design
is multidimensional. The hydrophobic substituent constant for the
adamantyl group indicates that the logP value of a compound with high
water solubility (logP ≪ 0) could be moved with an adamantyl-based
modification to a region that is more clinically useful. The steric bulk
of the adamantyl group can restrict or modulate intramolecular reac-
tivity; and impede the access of hydrolytic enzymes, thereby increasing
drug stability and plasma half-life. There are nearly forty adamantyl-
based compounds used to treat viral infections, neurodegenerative dis-
orders, acne vulgaris and type 2 diabetes mellitus. In almost all cases,
compound bearing an adamantyl are more lipophilic than the des-
adamantyl analogue [20].
was evaluated at λmax (260) by using Beer’s law (
ε
260 = 6600 cm
M )
-
4
and was found to be 1.65 × 10 M. The stock solution of compound (4)
was prepared in the ethanol–water mixture (1:1) and was optimized to
-
4
4.23 × 10 M. For cell line studies, DMEM (dulbecco’s modified eagle
medium), RPMI (Roswell park memorial institute) medium and (pen-
strep) MTT (3-[4,5-dimethylthiazol-2-yl]-2,5- diphenyltetrazolium bro-
mide were purchased from Thermo Fischer Scientific, USA and Sigma
Aldrich, USA, respectively.
2.2. Instrumentations
Melting point was recorded using a digital Gallenkamp (SANYO)
model MPD.BM 3.5 apparatus and are uncorrected. 1H and C NMR
spectra were determined in CDCl3 at 300 MHz and 75.4 MHz respec-
tively using a Bruker spectrophotometer. FTIR spectra were recorded on
an FTS 3000 MX spectrophotometer. Mass Spectra (EI, 70 eV) on a GC-
MS instrument Agilent technology, and elemental analyses were con-
ducted using a LECO-183 CHNS analyzer. A UV–visible spectropho-
tometer (Shimadzu-1800; TCC-240, Japan), with temperature controller
to maintain a required temperature of the sample in quartz cuvette, and
an automated Schott Gerate digital viscometer (Model; AVS 310) were
used for DNA binding experiments.
13
Adamantyl-1,3,4-oxadiazoles
thiadiazoles show antimicrobial, and anti-inflammatory activities
21]; adamantyl triazoles are selective inhibitors of 11β-hydroxysteroid
and
adamantylamino-1,3,4-
[
dehydrogenase type 1 [22] as are the thiazolidine derivatives with an
adamantyl group [23]. N,N-(1-diadamantane dicarboxami des) are po-
tential antitumor activity [24], 1-adamantane carboxylic acid hydra-
zides have antimicrobial activity [25] and N-arylsulsonyl-N-adamantyl
ureas show hypoglycemic activity [26]. A number of thiourea and
guanidine derivatives bearing adamantyl moiety were designed and
explored for their antimicrobial action against Gram-positive, Gram-
negative and a panel of drug-resistant Non-fermentative Gram-negative
Bacilli isolates from lung disease cystic fibrosis (CF) patients. The role of
different substituents on the antimicrobial activity was been determined
and a guanidine derivative bearing adamantane-1-carbonyl and 2-
bromo-4,6-difluouro-phenyl substituents (H-BDF), showed a potent
bactericidal activity against the strains tested, at levels higher than those
exhibited by the standard antibiotics tobramycin, ceftazimide and
meropenem. In addition, it was found to display low levels of cytotox-
icity against THP-I cells with a selective index (SI) > 8, pointing to its
potential for further development as a novel antibacterial drug [27].
Drug – DNA research could help to disclose a compound for its drug
nature. Non-covalent interaction of a drug with DNA via intercalation,
electrostatic interactions, hydrogen bonding, and van der Waals in-
teractions is generally considered more protective as, due to reversible
binding, healthy cells remain less intact with drug toxicity. In-vitro DNA
binding and cancer cell line studies for compound’s cytotoxicity on
cancerous and healthy cells are the useful approaches to diagnose the
candidacy of a compound for its anticancer activity [28,29].
2.3. Synthesis of 1-(Adamantane-1-carbonyl)-3-(1-naphthyl) thiourea
(4)
A freshly prepared solution of adamantane-1-carbonyl chloride (10
mmol) in dry acetone (50 ml) was added dropwise to a suspension of
ammonium thiocyanate (10 mmol) in dry acetone (30 ml) and the re-
action mixture was refluxed for 30 min under nitrogen. After cooling to
room temperature, a solution of the 1-naphthyl amine (10 mmol) in
acetone (10 ml) was added dropwise and the reaction mixture refluxed
for 4 h. On completion, as monitored by TLC, the reaction mixture was
poured into ice-water and the precipitated thiourea were recrystallized
from boiling ethanol.
2.4. Characterization data
◦
ꢀ 1
cm ): 3336 (NH), 3034 (Ar-CH),
Yeild 85%, mp 170 C. FT-IR (
ν
–
–
1
2909, 2849 (CH , CH), 1675, 1575, 1457, 1370 (C
2
S). H NMR (300
MHz, CDCl ): 8.70 (br s, 1H, NH, D O exchangeable); 8.10 (m, 1H, Ar),
3
δ
2
7.81 (m, 1H, Ar), 7.51 (m, 2H, Ar), 7.32 (m, 2H, Ar), 7.02 (m, 1H, Ar);
2.08 (s, 3H, adamantane-CH), 1.69 (s, 6H, adamantane-CH ), 1.58 (q,
2
1
3
Taking into account the aforesaid biological and synthetic signifi-
cance of thioureas on one hand and the multifunctional value of the
adamantyl group in drug design on the other, the endeavor of current
6H, adamantane-CH , J = 8.6 Hz); C NMR (75 MHz, CDCl ): 184.1
2
3
–
–
–
(C
–
S); 169.92 (C
O); 141.0 (C-NH);130.7, 131.2, 127.8, 126.9, 125.8,
124.9, 124.3, 121.4, 39.25, 38.69, 38.49, 36.44, 36.14, 28.05, 27.86,
2

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Scheme 1. Synthesis of 1-(Adamantane-1-carbonyl)-3-(1-naphthyl) thiourea (4).
2
7.78, (adamantane-C); Anal. Calcd for C22
H
24
N
2
OS (364.5): C, 72.49;
exchange correlation functional convincing that DFT could predict
molecular physicochemical properties and reaction barriers with accu-
H, 6.64; N, 7.69; S, 8.80 Found: C, 72.49; H, 6.64; N, 7.69; S, 8.80%.
racy [35]. Frontier molecular orbitals energies i.e; EHOMO. ELUMO
,
2
.5. X-ray crystallography
E
HOMO-ELUMO gap and molecular electrostatic potential (MEP) were also
computed at B3LYP/6-311G(d,p) level of theory.
The crystallographic data of the title compound (4) were collected on
a STOE IPDS II two-circle diffractometer using MoK
α
radiation (λ =
2.7.2. Molecular docking for DNA binding and urease enzyme inhibition
0
.71073 Å) at 173(2) K. The multi-scan absorption correction
Chemical computing Inc; Molecular Operating Environment MOE
2017, tool was employed to sketch and optimize the structure of 4 at a
MOPAC 7.0 level of theory. The structure was built on MOE molecular
builder and fetched to the MOE database after geometry relaxation. X-
ray crystallographic structure of DNA and urease enzyme possessing
PDB ID: 6CQ3 and PDB ID:1LPQ respectively were downloaded from
PDB [36,37] having resolution of 1.59 Ǻ and 1LPQ has a resolution of
3.4 Ǻ. Downloaded structures were protonated and optimized using the
protonate-3D menu of MOE modeling suite. Water molecules attached
with 6CQ3 and1LPQ were removed by means of sequence editor of MOE
panel. For the purpose of docking analysis, coordinates of 6CQ3 and
[
[
MULABS] applied data were processed by SHELX program packages
SHELXS97 and SHELXL97] for solving and refining the structure, and
ORTEP-3 and PLATON programs were used in drawings [30,31]. The
NH hydrogen atoms are located in a difference Fourier map and refined
freely. The remaining hydrogen atom positions were calculated
geometrically at distances of 0.95 Å (for aromatic CH) and 0.99 Å (for
2
CH ) and refined using a riding model by applying the constraints of
U
iso(H) = 1.2 X Ueq (C). Crystallographic data for the structure reported
herein have been deposited with the Cambridge Crystallographic Data
Centre (CCDC # 1965600).
1
LPQ were relaxed using an AMBER force field and a semiempirical PM3
2
.6. Hirshfeld surface calculations
approaches. Relaxed coordinates had a minimum energy and stable
conformation for best scoring function calculations. 4 was docked with
6CQ3 for DNA binding and with 1LPQ for urease inhibition studies. For
this purpose, the optimized structure was subjected to methodical mo-
lecular docking taking 6CQ3 and 1LPQ as a receptor at default param-
The Hirshfeld surface (HS) analysis was carried out by employing
Crystal Explorer program 17.5 [32]. The Hirshfeld surface distance from
the nearest nucleus inside and outside the surface was measured and
ꢀ 1
represented by d
tance was represented as dnorm. White, red and blue colors have been
selected for the visualization of dnorm
i
and d
e
, respectively, while a normalized contact dis-
eters with an RMS gradient of 0.01 kcal mol employing Site Finder to
find active sites of LPQ and 6CQ3. Several docking runs were allowed to
get final docking poses as perfect as possible. At each step of the simu-
lation, the energy of interaction of 4 with 6CQ3 and 1LPQ was deter-
mined. Rest of parameters were kept as a default [38].
.
2
2
.7. In silico methods for analysis
.7.1. Quantum chemical DFT studies
2.8. Experimental DNA binding procedure
Herein; quantum chemical DFT studies were performed in order to
get insight into erlecronic structure and properties of 4. Computational
modeling was performed using Gaussian 09 spackage installed in RCMS
integrated with Gauss View-05 to generated output strucutre, frontier
molecular orbitals and moelcular electrostatic potential energy surface
For DNA-binding investigations absorption titration experiments
◦
were run at 37 C and ct-DNA was added gradually from 10 up to 90 µM
-
4
into the fixed concentration (2.34 × 10 M) of 4. After each DNA
addition, the sample solution was kept at rest for five minutes to attain
◦
(
MESP) [33,34]. The compound was optimized by the hybrid B3LYP
the compound -DNA equilibrium and the required tmperature of 37 C.
functional with 6-311G(d,p) basis set. B3LYP is the first principal DFT
Viscosity experiments were run on digital viscometer to diagnose the
3

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
structure variation of DNA duplex in the presence of 4. By keeping
constant concentration of DNA solution, the small changes in the DNA
viscosity were observed through measurement of viscosity before and
Table 1
Experimental details.
Crystal data
after the addition of varying concentrations of 4 (10 up to 120 µM) at
Chemical formula
Mr
22 24 2
C H N OS
◦
364.49
3
7 C.
Crystal system, space group
Temperature (K)
Triclinic, P ꢀ 1
173(2)
a, b, c (Å)
6.7832 (5), 11.1810 (8), 13.666 (1)
2
2
.9. Bioassay
◦
α
, β, γ ( )
105.941 (6), 103.730 (6), 104.562 (6)
3
V (Å )
910.82 (13)
2
.9.1. Cytotoxicity of compound 4 on Huh-7 and Hek-293 cells
Z
Huh-7 and Hek-293 cells were obtained from Cell Culture Bank,
Radiation type
Mo K
α
0.19
◦
µ (mmꢀ
1
)
ASAB, NUST (Islamabad, Pakistan) and were stored at ꢀ 15 C. The Huh-
Crystal size (mm)
0.42 × 0.36 × 0.33
7
cell lines were grown in dulbecco’s modified eagle media (DMEM –
high glucose concentration) with 10% Fetal Bovine serum (FBS) and 1%
penicillin–streptomycin (pen-strep) Normal human embryonic kidney
cell line (Hek-293) was grown in RPMI high glucose media supple-
mented with 10% Fetal Bovine Serum (FBS) and 1% penciline-
Data collection
Diffractometer
STOE IPDS II two-circle diffractometer
Multi-scan MULABS (Spek, 2003; Blessing,
1995)
Absorption correction
◦
streptomycin. Cells were cultured at 37 C in the presence of 5% CO
2
Tmin, Tmax
0.924, 0.940
No. of measured, independent and
9181, 3389, 3068
and upon 80% confluency were split into the subsequent flasks till use.
The cells used for the cytotoxicity assay were 5th passage cells.
observed [I > 2
σ
(I)] reflections
R
int
0.036
0.608
sin θ/λ)max (Åꢀ
1
)
(
2
.9.2. Calorimetric MTT assay
For the MTT assay, Huh-7 were incubated in 96 well plates overnight
Refinement
with 10,000 cells per well. Five different concentrations (100, 200, 500,
50, 1000 nM) of the compound, dissolved in DMSO, were added in the
2
2
2
R[F > 2
σ
(F )], wR(F ), S
0.034, 0.091, 1.06
7
No. of reflections
No. of parameters
H-atom treatment
3389
243
wells in triplicates. DMSO was used as solvent control and 1% Triton-X
was as a positive control reference [39,40]. The cells were again incu-
H atoms treated by a mixture of
independent and constrained refinement
0.20, ꢀ 0.31
◦
bated along with 5% carbon dioxide at 37 C for 48 h. MTT reagent was
ꢀ 3
)
Δρ
max, Δ
ρmin (e Å
added into each well as well as in control well. The plates were then
further incubated for 3 h to completely achieve the purple formazan
precipitates. After aspiration of all solutions from the wells, formazan
was dissolved in 100 µL DMSO at room temperature and kept for 1 h in
the dark. The absorbance of each solution was recorded at 550 nm using
a microplate reader. The assay was repeated with Hek-293 cell line. All
assays were in three biological replicates at different time points.
From the MTT assay data of absorbances, percentage cell viability
and percentage cytotoxicity were calculated by using the following
equation;
Computer programs: X-AREA (Stoe & Cie, 2001), SHELXS97 (Sheldrick, 2008),
SHELXL97 (Sheldrick, 2008), XP (Sheldrick, 2008), SHELXL97.
peak at m/z 135 is derived from the adamantyl cation which on further
fragmentation results in weaker signals at m/z = 93, 80, 79, 67, 41 and
3
9. The other major fragments correspond to those of the N-McLafferty
rearrangement.
3.2. X-Ray structure
%
age of cell viability = (Asam ple-Ablank)/ (Acontrol-Ablank) × 100
1
-(Adamantane-1-carbonyl-3-(1-naphthyl)) thiourea (4) was further
characterized by single crystal X-ray diffraction study. The experimental
details are given in Table 1. The asymmetric unit of the title compound is
shown in Fig. 1 with the atom-numbering scheme. The dihedral angle
between the planar rings; A (C12—C17) and B (C11/C12/C17—C20) is
3
. Results and discussion
◦
A/B = 1.30(5) . So, they are co-planar. In the crystal structure, the
3
.1. Chemistry
intermolecular C
–
H⋅⋅⋅O hydrogen bonds (Table 2) link the molecules
2
into centrosymmetric dimers, enclosing R
2
(14) ring motifs (Fig. 2). On
H⋅⋅⋅O hydrogen bonds (Table 2)
The reaction sequence leading to the formation of title compound is
the other hand, the intramolecular N
–
depicted in Scheme 1. The starting material 1-adamantane carbonyl
chloride was obtained via the reaction of 1-adamantane carboxylic acid
with thionyl chloride [41]. A solution of adamantane-1-carbonyl chlo-
ride in dry acetone was treated with an equimolar quantity of potassium
thiocyanate in dry acetone to afford the adamantane-1-isothiocyanate as
intermediate. Treatment of the latter with an equimolar quantity of 1-
naphthylamine in dry acetone furnished the title thiourea derivative (4).
The 1-(Adamantane-1-carbonyl)-3-(1-naphthyl)thiourea thiourea
also enclose S(6) ring motifs (Fig. 2), in which they may be effective in
the stabilization of the structure.
3.3. Hirshfeld surface analysis
In order to visualize the intermolecular interactions in the crystal of
the title compound, a Hirshfeld surface (HS) analysis [42,43] was car-
ried out by using Crystal Explorer 17.5 [32]). In the HS plotted over
(
4) was characterized by FTIR absorptions at 3350, 3125 (free and
associated NH), 3031–3024 (Ar-CH), 2902–2926 (CH ), 2847–2850
CH), 1672 (carbonyl) and (2343) cm (thiocarbonyl) groups respec-
2
dnorm, the white surface indicates contacts with distances equal to the
ꢀ
1
(
sum of van der Waals radii, and the red and blue colours indicate dis-
tances shorter (in close contact) or longer (distinct contact) than the van
der Waals radii, respectively, Fig. 3 (left) [44]. The bright-red spots
appearing near S1 and hydrogen atom H26B indicate their roles as the
respective donors and/or acceptors; they also appear as blue and red
regions corresponding to positive and negative potentials on the HS
mapped over electrostatic potential [45,46] as shown in Fig. 3 (right).
The blue regions indicate the positive electrostatic potential (hydrogen-
bond donors), while the red regions indicate the negative electrostatic
1
tively. In the H NMR spectrum the characteristic signals of adamantyl
moiety: a 6H quartet at δ 1.75–1.79 (adamantane-CH ), a 6H, singlet at
.95–1.98 (adamantane-CH ) and a 3H, singlet around 2.08 (ada-
mantane-CH), besides singlets at δ 8.5–8.7 and 12.7–13.0 ppm were
2
1
2
1
3
observed for HN(1) and HN(2). In the C NMR characteristic signals at
for adamantyl moiety at δ 27.7, 36.1–36.4, 38.6–38.5 and 41.5–41.5 as
well those at δ 184.1 169.92 for thiocarbonyl carbons were observed.
Mass spectrum compound did not show the molecular ion peak; the base
4

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Fig. 1. The asymmetric unit of the title compound with the atom numbering scheme. Thermal ellipsoids are drawn at the 50% probability level.
potential (hydrogen-bond acceptors). The shape-index of the HS is a tool
Table 2
Hydrogen-bond geometry (Å, ).
to visualize the
triangles; if there are no adjacent red and/or blue triangles, then there
are no interactions. Fig. 3 (bottom) clearly suggests that there are
no interactions in (4).
The overall two-dimensional fingerprint plot, Fig. 4a, and those
delineated into H … H , H … C/C … H, H … S/S … H, H … O/ O … H, H
N/N … H, S … C/C … S, S … N/N … S, C … N/N … C and S … O/ O …
π
…
π
stacking by the presence of adjacent red and blue
◦
D—H⋅⋅⋅A
D—H
H⋅⋅⋅A
D⋅⋅⋅A
D—H⋅⋅⋅A
π … π
π
… π
N1—H1⋅⋅⋅O1
C26—H26B⋅⋅⋅S1
0.86 (2)
0.97
1.85 (2)
2.81
2.6003 (15)
3.7210 (15)
144.0 (17)
156
ii
Symmetry code: (ii) ꢀ x + 2, ꢀ y + 2, ꢀ z + 1.
…
Fig. 2. A partial packing diagram. The intramolecular N
H⋅⋅⋅O and intermolecular C H⋅⋅⋅O hydrogen bonds linking the molecules into centrosymmetric dimers are
– –
shown as dashed lines.
5

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Fig. 3. Left; View of the three-dimensional Hirshfeld surface of the title compound plotted over dnorm in the range of ꢀ 0.1210 to 1.4046 a.u, Right; View of the three-
dimensional Hirshfeld surface of the title compound plotted over electrostatic potential energy in the range ꢀ 0.0500 to 0.0500 a.u. using the STO-3 G basis set at the
Hartree–Fock level of theory. Hydrogen-bond donors and acceptors are shown as blue and red regions around the atoms corresponding to positive and negative
potentials, respectively, Bottom; Hirshfeld surface of the title compound plotted over shape-index. (For interpretation of the references to color in this figure legend,
the reader is referred to the web version of this article.) (For interpretation of the references to color in this figure legend, the reader is referred to the web version of
this article.)
S contacts [47] are illustrated in Fig. 4 b—j respectively, together with
their relative contributions to the Hirshfeld surface. The most important
interaction is H … H (Table 3) contributing 59.3% to the overall crystal
packing, which is reflected in Fig. 4b as the widely scattered points of
high density due to the large hydrogen content of the molecule with the
3.4. DFT structural analysis
Structural geometry of 4 has been simulated and optimized using
DFT/B3LYP to find geometric parameters (bond lengths, bond angles)
and electronic parameters (EHOMO, ELUMO, ΔE and MESP). The opti-
mized geometry with symmetric charge distribution on each individual
atom of 4 is displayed in Fig. 6 (left). The frontier molecular orbital
(FMOs) calculations were performed using quantum chemical DFT
which is a popular approach to describe distribution of electron den-
sities. The corresponding EHOMO, ELUMO values and HOMO-LUMO gap
(ΔE) are provided in Fig. 6 (right). It is apparent from the values that
electron transfer in 4 is more feasible due to smaller HOMO-LUMO gap.
To comprehend the distribution of isodensities, HOMO and LUMO sur-
faces were investigated revealing most distribution of isodensities on
heteratoms.
tip at de = di = 1.14 Å. In the absence of C
of characteristic wings resulting in the fingerprint plot delineated into H
C/C … H contacts, Fig. 4c, the 19.8% contribution to the HS are
viewed as pairs of spikes with the tips at at d = 2.72 Å and 2.88 Å
+ d
–
H …
π
interactions, the pairs
…
e
i
for big and small wings, respectively, arising from the H … C/C …
contacts (Table 3). The pair of characteristic wings resulting in the
H
fingerprint plots delineated into H … S/S … H, Fig. 4d, contacts with
1
0.1% contribution to the HS arises from the H … S/S … H contacts
(
Table 3) and is viewed as pair of spikes with the tips at d
The H … O/O … H contacts (Table 3) contributing 4.2% to the overall
crystal packing, are reflected in Fig. 4e with the tips at d = 2.62 Å.
+ d
The H … N/N … H contacts (Table 3), Fig. 4f, the 1.8% contribution to
the HS are viewed with the tips at d
e
+ d
i
= 2.67 Å.
e
i
Optimized geometry of 4 at B3LYP/6- 31G(d,p) level of theory was
also used to map the molecular electrostatic potential surfaces (MESP)
and is given in Fig. 6 (bottom). It depicts that negative potenial is
concentrated on oxygen, and nitrogen atoms which reflects elecron
transfer from O, and N (labelled with red contours). Dispersion of po-
tential for 4 ranges from ꢀ 0.0053 to 0.0053 esu indicating electrophilic
and nucleophilic centers in this range. Red color shows that –O and –N
centers contribute as nucleophilic regions, whereas grey color indicates
that –C and –H atoms contribute as electrophilic regions.
e
+ d = 3.18 Å. The S … C/C … S
i
(
Fig. 4g) and S … N/N … S (Fig. 4h) contacts (Table 3), contributing
1
.0% to the overall crystal packing, are viewed as pairs of spikes with the
+ d = 3.59 Å and d + d = 3.55 Å, respectively. Finally, the C
tips at d
e
i
e
i
…
N/N … C (Fig. 4i) and S … O/O … S (Fig. 4j) contacts (Table 3),
contributing 0.6% and 0.4%, respectively, to the overall crystal packing,
are reflected as the scattered points of low density.
The Hirshfeld surface representations with the function dnorm plotted
onto the surface are shown for the H … H, H … C/C … H and H … S/S …
H interactions in Fig. 5 a—c, respectively. The Hirshfeld surface analysis
confirms the importance of H-atom contacts in establishing the packing.
The large number of H … H, H … C/C … H and H … S/S … H in-
teractions suggest that van der Waals interactions and hydrogen
bonding play the major roles in the crystal packing [48].
The global chemical reactivity descriptors such as ionization poten-
tial (I = ꢀ EHOMO}), electron affinity (A = -ELUMO), hardness (
A)/ 2), softness (S = 1/ 2 ), chemical potential (µ = ꢀ (I + A)/ 2),
electronegativity ( / 2 )
η = (I ꢀ
η
2
χ
= (I + A)/ 2) and electrophilicity index (
ω
=
μ
η
are determined on the basis of the energies of frontier molecular orbital
[49] and given in Table 4. These global reactivity parameters indicate
that 4 is sufficiently stable due to high ionization energy. Significantly
high electronegativity reveals that 4 acts as electron withdrawing during
6

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Fig. 4. The full two-dimensional fingerprint plots for the title compound, showing (a) all interactions, and delineated into (b) H … H , (c) H … C/C … H, (d) H … S/S
and d values are the
…
H, (e) H … O/ O … H, (f) H … N/N … H, (g) S … C/C … S, (h) S … N/N … S, (i) C … N/N … C and (j) S … O/ O … S interactions. The d
i
e
closest internal and external distances (in Å) from given points on the Hirshfeld surface contacts.
7

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Table 3
bond formation with electron rich and electron donating DNA base pairs
and residues of urease enzymes. Smaller value of softness indicating
good polarizabilty measures and efficient overlap of electronic clouds of
4 with DNA base pairs and with the residues of urease enzyme.
Selected interatomic distances (Å).
i
S1⋅⋅⋅C20
3.2503 (15)
3.6378 (15)
3.5487 (14)
3.6220 (15)
2.77
C20⋅⋅⋅H26A
C26⋅⋅⋅H2
2.83
2.460 (18)
2.14
2.54
2.59
2.02
2.49
2.54
2.49
2.50
2.53
2.56
2.52
2.50
2.51
2.48
2.53
2.54
2.52
2.52
2.50
2.54
2.52
i
S1⋅⋅⋅C2
ii
S1⋅⋅⋅N2
H1⋅⋅⋅H13
i
iii
S1⋅⋅⋅C1
H1⋅⋅⋅H30B
3
.5. Molecular docking studies for DNA binding
S1⋅⋅⋅H20
S1⋅⋅⋅H2
H2⋅⋅⋅H26A
H2⋅⋅⋅H26B
H16⋅⋅⋅H18
H18⋅⋅⋅H24B
2.611 (18)
2.94
ii
S1⋅⋅⋅H22A
Molecular mechanism and the strength of interaction between 4 and
ii
v
v
S1⋅⋅⋅H26B
O1⋅⋅⋅H1
2.81
DNA was computationally investigated by molecular docking approach.
1.86 (2)
2.91
H18⋅⋅⋅H30A
H19⋅⋅⋅H29A
vi
Pose view analysis and conformations of 4 with lowest free energy is
H22B⋅⋅⋅O1
O1⋅⋅⋅H27A
O1⋅⋅⋅H27B
depicted in Fig. 7. Binding free energy (ΔG) and binding constant (K
b
)
2.49
H22A⋅⋅⋅H24A
H22A⋅⋅⋅H26B
H22B⋅⋅⋅H27A
H22B⋅⋅⋅H30B
H24A⋅⋅⋅H26B
H24A⋅⋅⋅H27B
H24B⋅⋅⋅H29A
H24B⋅⋅⋅H30A
H26A⋅⋅⋅H27B
H26A⋅⋅⋅H29B
H27A⋅⋅⋅H30B
H27B⋅⋅⋅H29B
H29A⋅⋅⋅H30A
2.91
values indicate that 4 interact with DNA spontaneously with efficient
binding propensity (Table 5). As far as interaction of 4 with DNA is
concerned, Fig. 7 (left) shows mixed mode of interaction. Planar naph-
thyl group intercalates between DNA base pairs DA (A6) and DC(B23)
and asymmetric structure turns to interact with minor groove of DNA
encompassing base-pairs DA(A5) and DG (B24). Hydrophobic in-
teractions of 4 with DNA are indicated with dotted lines in lig plot, Fig. 7
(right).
iii
O1⋅⋅⋅H22B
O1⋅⋅⋅H30B
2.72
iii
2.73
iv
N1⋅⋅⋅C14
3.450 (2)
2.54
iv
N1⋅⋅⋅H13
N2⋅⋅⋅H26A
N2⋅⋅⋅H26B
C1⋅⋅⋅H20
C2⋅⋅⋅H1
2.91
2.62
2.89
2.361 (19)
2.92
i
i
C11⋅⋅⋅H26A
C13⋅⋅⋅H1
2.63 (2)
2.96
i
C18⋅⋅⋅H25
3.6. Molecular docking studies for urease enzyme inhibition
C19⋅⋅⋅H26A
2.99
Symmetry codes: (i) ꢀ x + 1, ꢀ y + 2, ꢀ z + 1; (ii) ꢀ x + 2, ꢀ y + 2, ꢀ z + 1; (iii) ꢀ
x + 1, ꢀ y + 1, ꢀ z + 1; (iv) × + 1, y, z; (v) × ,y + 1, z + 1; (vi) × + 1, y + 1, z + 1.
Molecular docking was also run to theoretically examine the possible
interactions that compound 4 could execute with urease enzyme for
urease inhibition. For that purpose, free docking conformation of 4
within active site of urease, lowest energy docking pose of 4 within
Fig. 5. The Hirshfeld surface representations with the function dnorm plotted onto the surface for (Left) H … H, (Right) H … C/C … H and (Bottom) H … S/S …
interactions.
H
8

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Fig. 6. Left; The optimized geometry of 4 calculated at the B
3
LYP/6-31G (d,p) level of theory, Right; HOMO, LUMO, EHOMO, ELUMO and band gap (ΔE) for the title
compound, Bottom; Molecular electrostatic potential surfaces (MESP) of 4 mapped at B3LYP/6-31G (d,p) level of theory.
Table 4
Global chemical reactivity descriptors obtained from B3LYP with basis set 6-31G (d,p).
Compound
E
HOMO/eV
E
LUMO/eV
I /eV
3.90
A /eV
2.25
η
/eV
S /eV
μ
/eV
χ
/eV
ω
/eV
4
ꢀ 3.90
ꢀ 2.25
ꢀ 0.825
ꢀ 0.606
ꢀ 3.075
3.075
ꢀ 5.73
binding pocket of urease structure and lig plot are shown in Fig. 8. Pose-
could be related to the presence of planner naphthyl group and lone
pairs present on oxygen of carbonyl group and on sulfur of thiourea
moiety. As the naphthyl group has planner nature so it can intercalate
between the base pairs of DNA and hydrogen bonding arises between the
lone pairs of oxygen and sulfur with the partial positively charged hy-
view analysis and computationally calculated binding parameters (ΔG,
K ) shows that 4 fits well into binding pocket of urease to develop in-
b
teractions and inhibit its active site, Fig. 8 (left). Lig plot is indicative of
urease pocket interactions with 4 in two forms; (1) hydrophobic in-
teractions of 4 with amino acid residues of urease enzyme pocket, (2)
arene-arene interactions of Methionine (Met 475) with aromatic ring of
drogens of –NH
2
and –NH groups of DNA bases.
The strength of binding tendency was calculated in terms of binding
Ao
Aꢀ Ao
ε
G
4
, Fig. 8 (right). The binding parameters in Table 5 indicate that in-
constant (K ) by using Bnisi- Hildebrand equation; {
=
+
b
ε
Hꢀ G
ꢀ εG
teractions of 4 with DNA and urease enzymes are spontaneous in nature.
Docking results reveal that compounds 4 binds effectively with urease
enzyme, however these findings may further be verified experimentally.
ε
G
1
}
, where, A
o
, A and
ε
G
and H-G represent absorption and
ε
ε
Hꢀ Gꢀ
ε
G
Kb[DNA]
molar extinction coefficient of 4 and 4 – DNA bound complex, respec-
tively [51]. While, to determine reaction spontaneity, free energy
change was calculated by using Vant Hoff’s equation {ΔG = -RT lnK
The binding constant (K ) was obtained from intercept to slope ratio of a
plot of A /A-A vs. 1/[DNA], Fig. 9 (left). The calculated binding pa-
rameters are provided in Table 5. The evaluated binding constant (K
b
}.
3
.7. UV–visible spectroscopic and viscometric studies for DNA binding
UV–visible spectroscopy is one of the most reliable, accessible, and
b
o
o
b
applicable technique to describe a compound for its binding with DNA
49–51]. The absorption spectrum of 4 was recorded in the absence and
ꢀ 1
4
/
M
) was 10 order of magnitude and the value was found comparable
[
with the K
b
values of commonly used drugs i.e., Ciprofloxacin (1.34 ×
in the presence of increasing concentrations of DNA and the variations in
the spectral response were monitored, Fig. 9 (left). The absorption band
of the compound was affected and a gradual decrease in the absorption
peak intensity was observed in the presence of an increasing concen-
4
4
4
1
0 ) [52], Lumazine (1.74 × 10 ) [53], Valrubicin (1.75 × 10 ) [54],
4
and Citral (1.12 × 10 ) [55]. The negative value of free energy change
indicated that the compound’s interaction with DNA take place in
spontaneous mode [56]. A comparison of DNA binding of 4 with some
reported thiourea derivatives is also provided in Table 6, which further
pointed towards significant binding of 4 with DNA [43,50,51,57–61].
The interactional modes, as claimed through various techniques,
could be further verified through the measurement of DNA viscosity in
the presence of compound’s concentrations [62]. The viscosity of DNA
increases when base pairs of DNA duplex accommodate the drug
tration of DNA. This hypochromic effect was evaluated by using the
Afreeꢀ Abound
equation; {H% =
× 100}, where Afree and Abound are the
Afree
absorbance of 4 and 4 – DNA complex, respectively. The percent
hypochromism was evaluated to be 71.5% along with a prominent red
shift of 5.1 nm. Spectral findings indicated that 4 could bind with DNA
via intercalation [49,51]. Further, the binding tendency of 4 with DNA
9

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Fig. 7. Left; Molecular docked 4 – DNA complex, Right; Lig plot of 4 with DNA base pairs calculated at PM3 semi-empirical level of theory.
Table 5
Binding parameters calculated from molecular docking and UV–visible spectroscopic data.
Complex
Molecular docking
UV–visible spectroscopy
ꢀ
1
ꢀ ΔG / kJmolꢀ 1
ꢀ 1
ꢀ ΔG / kJmolꢀ 1
K
b
/ M
b
K / M
3
3
4
4
4
- DNA
2.22 × 10
5.06 × 10
18.97
21.21
1.56 × 10
24.88
- urease enzyme
–
–
molecule. This rise in viscosity is due to the intercalative mode of in-
teractions between the compound and DNA [63]. The decrease in vis-
cosity of DNA is observed when a compound bind with DNA through
groove binding or electrostatic interactions due to the twisting of DNA
helix [64]. If the viscosity of DNA solution rises up to a certain limit and
then becomes constant, then it verifies the mixed mode of interactions
carcinoma cell lines (Huh-7) as well as normal human embryonic kidney
cell lines (Hek-293) i.e. non-cancerous/normal cells.
Cell cytotoxicity data of the compound is given in Fig. 10 and the
values of percentage of cell viability and cell cytotoxicity for each
compound are given in Table 7. All statistical analyses were performed
using the GraphPad Prism 5 software and a p value < 0.05 was
considered as statistically significant.
(
intercalation along with groove binding or electrostatic interactions)
65].
o
In viscosity experiments, initially, DNA viscosity ( ) of was
[
4 showed dose-dependent cytotoxic activity against both cancerous
hepatocellular carcinoma cell line as well as the normal human em-
bryonic kidney cell line when tested for concentration ranges of
100–1000 nM. Percentage cell viability remained greater than 50% at
concentrations between 100 and 750 nM (p < 0.001) of the compound at
non-cancerous Hek-293 cell line. The cell viability at the cancerous Huh-
7 cell line however remained less than 50% even at the lowest drug
concentration i.e., 100 nM, indicating that the compound 4 is selectively
more cytotoxic towards cancer cells, even at lower concentration, than
normal cells. The results suggest significant anticancer potential of 4
against hepatocellular carcinoma cell lines at the tested concentrations
and conditions. Further, in comparison to our previously reported work
on three aroylthiourea derivatives of ciprofloxacin drug ATU-1, ATU-2
& ATU-3, the values of % cell viability and % cytotoxicity of 4 at 1000
nM concentration indicated that the current studied compound has
much improved anticancer potency than that of ATU-1 — ATU-3 where
% cell viability and % cytotoxicity was found to be 22.45, 32.45, 40.15
η
measured and then varying concentrations of 4 were gradually added
into DNA fixed concentration and the changes in DNA viscosity was
1
/3
measured as
η
. The cube root of relative viscosity (
η
/
η
o
)
is plotted vs.
[
Compound/[DNA] ratio and is shown in Fig. 9 (bottom). The viscosity
of DNA solution increased upon gradual addition of 4. However, after
the addition of 90 µM of 4, further addition till 120 µM did not showed
any considerable change; hence verified the mixed binding mode [65].
3
.8. Cell line studies for anticancer/cytotoxic activity
The effects of a compound on cell line activity is one of the important
in-vitro studies that could help to correlate the data obtained by various
theoretical and experimental studies and to further help in determining
anticancer potency of a compound for targeted cancer. The percentage
cell viability and cytotoxicity activity of tested compound 4 was eval-
uated with the help of MTT assay by using various concentrations of 4.
Anticancer potential of 4 was checked against human hepatocellular
and 77.55, 67.55, 59.85, respectively, at 100 g/mL [51].
μ
1
0

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Fig. 8. Left; Molecular docked complexes of 4 with urease enzyme, Right; Lig plot of 4 with residues of urease enzyme calculated at PM3 semi-empirical level
of theory.
Fig. 9. Left; UV–visible spectrum of 4 and spectral variations upon addition of DNA concentrations from 10 ꢀ 90 at 37 ◦C, Right; plot of A
0
0
/A-A vs. 1/[DNA] for the
determination of binding constant, Bottom; Plot of relative specific viscosity vs. compound to DNA concentration ratio for the verification of binding modes.
1
1

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
Table 6
Comparison DNA binding of 4 with some related thiourea derivatives.
Thiourea derivatives
DNA binding studies
/M
References
-ΔG / kJmolꢀ
1
K
b
1
-(adamantane-1-carbonyl-3-(1-naphthyl)) thiourea (4)
Table 5
Table 5
24.89
Present study
4
N-(4-(1H-benzo[d]imidazol-2-yl)phenyl)-3-benzoyl thiourea (TUBC)
N-((2-(benzo [4,5] imidazo [1,2-c]quinazolin-6 yl)
2.31 × 10
43
50
4
2.9 × 10
26.49
phenyl)carbamothioyl) heptanamide—quinolinyl-thiourea hybrid (3)
Aroylthiourea derivatives of ciprofloxacin drug (ATU-1 — ATU-3)
4
7.93, 6.22, 1.66 × 10
29.07, 28.44,
51
2
5.04
4
Ru(II)-based complexes with N-(acyl)-N’,N’-(disubstituted)thiourea ligands
Palladium(II) complex with thiourea (1–3)
0.8–1.8 × 10
27.13–28.45
–
–
57
58
59
4
8.6, 6.7, 7.3 × 10
′
′
′
4
3
Cu(II) and Ni(II) metallamacrocycles formed from 3, 3, 3 , 3 tetrabenzyl-1,1 -aroylbis(thiourea) ligands(H
2
L1, 1,
1.52, 1.98, 2.38 × 10
2
)
3
1
-(2-florobenzoyl)-3-(2-chloro,4-ferrocenylphenyl) thiourea (2F)
2.92 × 10
19.76
60
61
Cu(I) thiourea derivatives with triphenylphosphine(I
1
— I
3
)
8.19, 5.65, 5.31 × 10
22.33, 21.41, 15.54
pointed towards mixed mode binding between 4 - DNA and the same
was verified in viscometric studies; hence computational and experi-
mental findings complemented each other. Moreover, the results of the
′
cytotoxicity assay suggest that compound 4 s antiproliferative activity
seems to be selective for tumor cells as much less toxicity against normal
cells was observed even at comparatively higher concentrations. Thus,
this study supports the use of this compound in further preclinical
studies for evaluation of its effects on malignant tumors.
Declaration of Competing Interest
The authors would like to declare that there is no established conflict
of interest.
Acknowledgments
Fig. 10. Percentage viability of tested compound 4 at increasing concentration
on Hek-293 and Huh-7 cell line, respectively. DMSO was used as solvent control
and 1% Triton-X-100 was used as positive reference control.
The contributions of all authors’ organizations for the facilitation of
this collaborative research is highly acknowledged.
Table 7
References
Cell line activity data of 4 for normal human embryonic kidney cell line (Hek-
[1] A. Saeed, M.N. Mustafa, M. Zain-ul-Abideen, G. Shabir, M.F. Erben, U. Fl o¨ rke,
Current developments in chemistry, coordination, structure and biological aspects
of 1-(acyl/aroyl)-3- (substituted)thioureas: advances Continue … , J. Sulfur Chem.
40 (3) (2019).
2
93) and cancerous human hepatocellular carcinoma cell line (Huh-7).
Conc.
nM)
Hek-293
Huh-7
(
%
Cell
%
% Cell
%
[
2] F.A. Larik, A. Saeed, P.A. Channar, U. Muqadar, Q. Abbas, M. Hassan, S.-Y. Seo,
M. Bolte, Design, Synthesis, kinetic mechanism and molecular docking studies of
novel 1-pentanoyl-3-arylthioureas as inhibitors of mushroom tyrosinase and free
radical scavengers, Eur. J. Med. Chem. 141 (2017) 273–281.
viability
Cytotoxicity
viability
Cytotoxicity
1
2
5
7
1
00
75.2 ± 1.11
69.6 ± 1.2
62.3 ± 1.6
51.3 ± 1.6
40 ± 2
24.8
30.4
37.7
48.7
60
42.9 ± 0.82
41.8 ± 0.37
30.3 ± 0.7
57.1
58.2
69.6
74.6
81.04
00
[
3] A. Saeed, S.-U. Rehman, P.A. Channar, F.A. Larik, Q. Abbas, M. Hassan, H. Raza,
U. Fl o¨ rke, S.-Y. Seo, Long chain 1-acyl-3-arylthioureas as jack bean urease
inhibitors, synthesis, kinetic mechanism and molecular docking studies, J. Taiwan
Inst. Chem. Eng. 77 (2017) 54–63.
00
50
25.4 ± 0.56
18.96 ± 0.23
000
[
[
[
4] L.-P. Duan, J. Xue, L.-L. Xu, H.-B. Zhang, Synthesis 1-acyl-3-(2’-aminophenyl)
thioureas as anti-intestinal nematode prodrugs, Molecules 15 (10) (2010)
4
. Conclusions
6
941–6947.
5] M. Struga, S. Rosolowski, J. Kossakowski, J. Stefanska, Synthesis and
microbiological activity of thiourea derivatives of azatricycloundec-8-ene-3,5-
dione, Arch. Pharm. Res. 33 (1) (2010) 47–54.
Novel adamantane-naphthyl thiourea conjugate (4), was synthesized
in excellent yield by a one pot method from prepared adamantane-1-
carbonyl chloride and characterized spectroscopically. The X-ray anal-
ysis designates it to belonging to triclinic system P-1 space group and
planarity of naphthyl group. In the crystal structure, the intermolecular
6] A. Saeed, H. Rafique, A. Hammed, S. Rasheed, Synthesis and antibacterial activity
of some new 1-aroyl-3-(substituted-2-benzothiazolyl)thioureas, Pharm. Chem. J.
1
8 (5) (2009) 152–158.
[7] A. Saeed, U. Shaheen, A. Hameed, S.H. Naqvi, Synthesis, characterization and
antimicrobial activity of some new 1-(fluorobenzoyl)-3-(fluorophenyl)thioureas,
J. Fluorine Chem. 130 (11) (2009) 1028–1034.
C
–
H⋅⋅⋅O hydrogen bonds link the molecules into centrosymmetric di-
2
mers, enclosing R
2
(14) ring motifs. On the other hand, the intra-
[
8] S. Karaku s¸ , S. Rollas, Synthesis and antituberculosis activity of new N-phenyl-N’-4-
molecular N
–
H⋅⋅⋅O hydrogen bonds also enclose S(6) ring motifs. The
(5-alkyl/arylamino-1,3,4-thiadiazole-2-yl)phenylthioureas, Il Farmaco 57 (7)
(
2002) 577–581.
9] A. Liav, S.K. Angala, P.J. Brennan, M. Jackson, N-D-Aldopentofuranosyl-N -p-
isoamyloxy)phenyl thioureas, (isoamyloxy)phenyl] thiourea derivatives: Potential
Hirshfeld surface analysis of the crystal structure indicates that the most
important contributions for the crystal packing are from H … H (59.3%),
H … C/C … H (19.8%) and H … S/S … H (10.1%) interactions.
Hydrogen bonding and van der Waals interactions are the dominant
interactions in the crystal packing. DFT studies revealed efficient elec-
tron transfer, higher stability and electron withdrawing capacity of 4
while interacting with DNA base pairs and residues of urease enzyme.
DNA binding studies by molecular docking and UV–visible spectroscopy
′
[
(
anti-TB therapeutic agents, Bioorg. Med. Chem. Lett. 18 (8) (2008) 2649–2651.
[
10] D. Sriram, P. Yogeeswari, M. Dinakaran, R. Thirumurugan, Antimycobacterial
activity of novel 1-(5-cyclobutyl-1,3-oxazol-2-yl)-3(sub)phenyl/ pyridyl thiourea
compounds endowed with high activity toward multidrug-resistant
Mycobacterium tuberculosis, J. Antimicrob. Chemother. 59(6) (2007) 1194-1196.
[11] N. Walchshofer, I. Delabre-Defayolle, J. Paris, A.F. Petavy, In vivo morphological
damage induced by a new benzimidazole prodrug in Echinococcus multilocularis
metacestodes, J. Pharm. Sci. 79 (7) (1990) 606–608.
1
2

N. Arshad et al.
Bioorganic Chemistry 109 (2021) 104707
[
[
[
[
12] A. Mishra, K. Srivastava, R. Tripathi, S. Puri, S. Batra, Search for new
pharmacophores for antimalarial activity. Part III: synthesis and bioevaluation of
new 6-thioureido-4-anilinoquinazolines, Eur. J. Med. Chem. 44 (11) (2009)
[37] https://www.rcsb.org/structure/6CQ3.
[38] https://www.rcsb.org/structure/1LPQ.
[39] F. Perveen, N. Arshad, R. Qureshi, J. Nowsherwan, A. Sultan, B. Nosheen,
H. Rafique, Electrochemical, spectroscopic and theoretical monitoring of
anthracyclines’ interactions with DNA and ascorbic acid by adopting two routes:
cancer cell line studies, PLoS ONE 13 (10) (2018), e0205764.
4
404–4412.
˙
13] P. Çıkla, G. Küçükgüzel, I. Küçükgüzel, S. Rollas, E. De Clercq, C. Pannecouque,
¨
G. Andrei, R. Snoeck, F. S¸ ahin, O. Bayrak, Synthesis and evaluation of antiviral,
antitubercular and anticancer activities of some novel thioureas derived from 4-
aminobenzohydrazide hydrazones, Marmara Pharm. J. 14 (1) (2010) 13–20.
14] H.-G. Park, J.-Y. Choi, S.-H. Choi, M.-K. Park, J. Lee, Y.-G. Suh, H. Cho, U. Oh,
J. Lee, S.-U. Kang, N-4-Substitutedbenzyl- N’-tert-butylbenzyl thioureas as
vanilloid receptor ligands: investigation on the role of methanesulfonamido group
in antagonistic activity, Bioorg Med. Chem. Lett. 14 (3) (2004) 787–791.
15] R.B. Patel, K.H. Chikhalia, C. Pannecouque, E.d. Clercq, Synthesis of novel PETT
analogues: 3,4-dimethoxy phenyl ethyl-1,3,5-triazinyl thiourea derivatives and
their antibacterial and anti-HIV studies, J. Braz. Chem. Soc. 18 (2) (2007)
[40] M.R. Mozafari, C. Reed, C. Rostron, Cytotoxicity evaluation of anionic
nanoliposomes and nanolipoplexes prepared by the heating method without
employing volatile solvents and detergents, Die Pharmazie- Int J. Pharm. Sci. Res.
62 (3) (2007) 205–209.
˙
[41] P. Prochor, Z.A. Mierzejewska, Influence of the surface roughness of PEEK GRF30
and Ti6Al4V SLM on the viability of primary human osteoblasts determined by the
MTT test, Materials 12 (24) (2019) 4189.
[42] F.L. Hirshfeld, Bonded-atom fragments for describing molecular charge densities,
Theor. Chim. Acta 44 (2) (1977) 129–138.
3
12–321.
[43] N. Arshad, M. Rafiq, R. Ujan, A. Saeed, S.I. Farooqi, F. Perveen, P.A. Channar,
S. Ashraf, Q. Abbas, A. Ahmed, T. Hokelek, M. Kauri, J.P. Jasinski, Synthesis, X-ray
crystal structure elucidation and Hirshfeld surface analysis of N-((4-(1H-benzo[d]
imidazole-2-yl) phenyl) carbamothioyl) benzamide: Investigations for elastase
inhibition, antioxidant and DNA binding potentials for biological applications, RSC
Adv. 10 (2020) 20837.
[
[
[
[
[
16] C. Sun, H. Huang, M. Feng, X. Shi, X. Zhang, P. Zhou, A novel class of potent
influenza virus inhibitors: polysubstituted acylthiourea and its fused heterocycle
derivatives, Bioorg. Med. Chem. Lett. 16 (1) (2006) 162–166.
17] A. Paul, L.C. Harrington, J.G. Scott, Evaluation of novel insecticides for control of
the dengue vector, Aedes aegypti (Diptera: Culicidae), J. Med. Entomol. 43 (1)
(
2006) 55–60.
[44] M.A. Spackman, D. Jayatilaka, Hirshfeld surface analysis, CrystEngComm 11 (1)
(2009) 19–32.
18] K. Ramadas, G. Suresh, N. Janarthanan, S. Masilamani, Antifungal activity of 1,3-
disubstituted symmetrical and unsymmetrical thioureas, Pestic. Sci. 52 (2) (1998)
[45] P. Venkatesan, S. Thamotharan, A. Ilangovan, H. Liang, T. Sundius, Crystal
structure, Hirshfeld surfaces and DFT computation of NLO active (2E)-2-
(ethoxycarbonyl)-3-(1-methoxy-1-oxo-3-phenylpropan-2-yl)amino prop-2-enoic
acid, Spectrochim. Acta Part A 153 (2016) 625–636.
1
45–151.
19] S.-Y. Ke, S.-J. Xue, Synthesis and herbicidal activity of N-(o-fluorophenoxyacetyl)
thioureas derivatives and related fused heterocyclic compounds, ARKIVOC 10
(
2006) 63–68.
[46] M.A. Spackman, J.J. McKinnon, D. Jayatilaka, Electrostatic potentials mapped on
Hirshfeld surfaces provide direct insight into intermolecular interactions in crystals
CrystEngComm. 10(4) (2008) 377-388.
20] S. Kumar, V. Awasthi, J. Kanwar, Influence of growth regulators and nitrogenous
compounds on in vitro bulblet formation and growth in oriental lily, Hort. Sci.
(
Prague) 34 (2) (2007) 77–83.
[47] D. Jayatilaka, D. Grimwood, A. Lee, A. Lemay, A. Russel, C. Taylor, S. Wolff, P.
Cassam-Chenai, A. Whitton, TONTO-a system for computational chemistry, 2005.
[48] J.J. McKinnon, D. Jayatilaka, M.A. Spackman, Towards quantitative analysis of
intermolecular interactions with Hirshfeld surfaces, Chem. Commun. 37 (2007)
3814–3816.
[
[
21] J. Liu, D. Obando, V. Liao, T. Lifa, R. Codd, The many faces of the adamantyl group
in drug design, Eur. J. Med. Chem. 46 (6) (2011) 1949–1963.
22] A.A. Kadi, N.R. El-Brollosy, O.A. Al-Deeb, E.E. Habib, T.M. Ibrahim, A.A. El-Emam,
Synthesis, antimicrobial, and anti-inflammatory activities of novel 3-(1-
adamantyl)-5-substituted-1,3,4-oxadiazoles and 2-(1-adamantylamino)-5-
substituted-1,3,4-thiadiazoles, Eur. J. Med. Chem. 42 (2) (2007) 235–242.
23] S. Olson, S.D. Aster, K. Brown, L. Carbin, D.W. Graham, A. Hermanowski-Vosatka,
C.B. LeGrand, S.S. Mundt, M.A. Robbins, J.M. Schaeffer, Adamantyl triazoles as
selective inhibitors of 11β-hydroxysteroid dehydrogenase type 1, Bioorg. Med.
Chem. Lett. 15 (19) (2005) 4359–4362.
[49] S.I. Farooqi, N. Arshad, P.A. Channar, F. Perveen, A. Saeed, F.A. Larik, A. Javed,
Synthesis, theoretical, spectroscopic and electrochemical DNA binding
investigations of 1, 3, 4-thiadiazole derivatives of ibuprofen and ciprofloxacin:
cancer cell line studies, J. Photochem. Photobiol B: Biol 189 (2018) 104–118.
[50] M. Hussain, N. Arshad, R. Ujan, A. Saeed, P.A. Channar, F. Perveen, F.A. Larik, S.
I. Farooqi, Z. Hussain, T. H o¨ kelek, M. Kaur, J.P. Jasinski, Synthesis, structure
elucidation and surface analysis of a new single crystal N-((2-(benzo [4,5]imidazo
[1,2-c]quinazolin-6-yl)phenyl) carbamothioyl)heptanamide: Theoretical and
experimental DNA binding studies, J. Mol. Struct. 1205 (2020), 127496.
[51] S.I. Farooqi, N. Arshad, F. Perveen, P.A. Channar, A. Saeed, F.A. Larik, A. Javeed,
Aroylthiourea derivatives of ciprofloxacin drug as DNA binder: Theoretical,
spectroscopic and electrochemical studies along with cytotoxicity assessment,
Arch. Biochem. Biophys. 666 (2019) 83–98.
[
[
24] S.W. Kwon, S.K. Kang, J.H. Lee, J.H. Bok, C.H. Kim, S. Dal Rhee, W.H. Jung, H.
Y. Kim, M.A. Bae, J.S. Song, Synthesis and 11β-hydroxysteroid dehydrogenase 1
inhibition of thiazolidine derivatives with an adamantyl group, Bioorg. Med.
Chem. Lett. 21 (1) (2011) 435–439.
[
[
[
[
[
25] P. Ficarra, R. Ficarra, A. Tommasini, G. Fenech, Compounds with potential
antitumor activity. II. N,N’-(1-diadamantane dicarboxamides), Il Farmaco;
edizione scientifica, 38(6) (1983) 418-424.
26] R. Ficarra, P. Ficarra, A. Tommasini, G. Fenech, F. Pizzimenti, G. Bisignano, 1-
Adamantane carboxylic acid hydrazides with presumed antimicrobial activity,
Boll. Chim. Farm 123 (1984) 317–321.
[52] H. Nawaz, S. Rauf, K. Akhtar, A.M. Khalid, Electrochemical DNA biosensor for the
study of ciprofloxacin–DNA interaction, Anal. Biochem. 354 (1) (2006) 28–34.
[53] M. Ibrahim, I. Shehatta, A. Al-Nayeli, Voltammetric studies of the interaction of
lumazine with cyclodextrins and DNA, J. Pharm. Biomed. Anal. 28 (2) (2002)
217–225.
27] A. Saeed, A. Bosch, M. Bettiol, D.L.N. Gonz ´a lez, M.F. Erben, Y. Lamberti, Novel
guanidine compound against multidrug-resistant cystic fibrosis-associated
bacterial species, Molecules 23 (5) (2018) 1158.
[54] R. Hajian, P. Hossaini, Z. Mehrayin, P.M. Woi, N. Shams, DNA-binding studies of
valrubicin as a chemotherapy drug using spectroscopy and electrochemical
techniques, J. Pharm. Anal. 7 (3) (2017) 176–180.
28] N. Kumar, N. Lal, V. Nemaysh, P.M. Luthra, Design, synthesis, DNA binding studies
and evaluation of anticancer potential of novel substituted biscarbazole derivatives
against human glioma U87 MG cell line, Bioorg. Chem. 100 (2020), 103911.
29] K.P. Rakesh, H.K. Kumara, H.M. Manukumar, D.C. Gowda, Anticancer and DNA
binding studies of potential amino acids based quinazolinone analogs: Synthesis,
SAR and molecular docking, Bioorg. Chem. 87 (2019) 252–264.
[55] M.F. Alam, S. Varshney, M.A. Khan, A.A. Laskar, H. Younus, In vitro DNA binding
studies of therapeutic and prophylactic drug citral, Int. J. Biol. Macromol. 113
(2018) 300–308.
[56] E. Movahedi, A.R. Rezvani, New silver (I) complex with diazafluorene based
ligand: synthesis, characterization, investigation of in vitro DNA binding and
antimicrobial studies, J. Mol. Struct. 1139 (2017) 407–417.
[
[
30] G.M. Sheldrick, A short history of SHELX, Acta Cryst. 64 (1) (2008) 112–122.
31] L.J. Farrugia, WinGX and ORTEP for Windows: an update, J. Appl. Crystallogr. 45
(
4) (2012) 849–854.
[57] R.S. Correa, K.M. de Oliveira, F.G. Delolo, A. Alvarez, R. Mocelo, A.M. Plutin, M.
R. Cominetti, E.E. Castellano, A.A. Batista, Ru(II)-based complexes with N-(acyl)-
N’, N’-(disubstituted)thiourea ligands: synthesis, characterization, BSA- and DNA-
binding studies of new cytotoxic agents against lung and prostate tumour cells,
J. Inorg. Chem. 150 (2015) 63–71.
[
[
32] A.L. Spek, Acta Crystallogr, D. Sect, Structure validation in chemical
crystallography, Biol. Crystallogr. 65 (2) (2009) 148–155.
33] M. Turner, J. McKinnon, S. Wolff, D. Grimwood, P. Spackman, D. Jayatilaka, M.
Spackman, CrystalExplorer17, Google Scholar There is no corresponding record for
this reference (2017).
[58] C.V. Barra, F.V. Rocha, L. Morel, A. Gautier, S.S. Garrido, A.E. Mauro, R.C.G. Frem,
A.V.G. Netto, DNA binding, topoisomerase inhibition and cytotoxicity of palladium
(II) complexes with 1,10-phenanthroline and thioureas, Inorganica Chim. Acta 446
(2016) 54–60.
[
34] M.J. Frisch, G.W. Trucks, H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R. Cheeseman,
G. Scalmani, V. Barone, G.A. Petersson, H. Nakatsuji, X. Li, M. Caricato, A.
Marenich, J. Bloino, B.G. Janesko, R. Gomperts, B. Mennucci, H.P. Hratchian, J.V.
Ortiz, A.F. Izmaylov, J.L. Sonnenberg, D. Williams-Young, F. Ding, F. Lipparini, F.
Egidi, J. Goings, B. Peng, A. Petrone, T. Henderson, D. Ranasinghe, V.G.
Zakrzewski, J. Gao, N. Rega, G. Zheng, W. Liang, M. Hada M. Ehara, K. Toyota, R.
Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda, O. Kitao, H. Nakai, T.
Vreven, K. Throssell, J.A. Montgomery Jr., J.E. Peralta, F. Ogliaro, M. Bearpark, J.
J. Heyd, E. Brothers, K.N. Kudin, V.N. Staroverov, T. Keith, R. Kobayashi, J.
Normand, K. Raghavachari, A. Rendell, J.C. Burant, S.S. Iyengar, J. Tomasi, M.
Cossi, J.M. Millam, M. Klene, C. Adamo, R. Cammi, J.W. Ochterski, R.L. Martin, K.
Morokuma, O. Farkas, J.B. Foresman, D.J. Fox, Gaussian 09, Rev. C.01, Gaussian,
Inc., Wallingford CT, 2010.
35] R. Dennington, T. Keith, J. Millam, GaussView 05, Semichem Inc., 2009.
36] M.N. Ahmed, K.A. Yasin, K. Ayub, T. Mahmood, M.N. Tahir, B.A. Khan, M. Hafeez,
M. Ahmed, Click one pot synthesis, spectral analyses, crystal structures, DFT
studies and brine shrimp cytotoxicity assay of twonewly synthesized 1,4,5-trisub-
stituted 1,2,3-triazoles, J. Mol. Struct. 1106 (2016) 430–439.
[59] N. Selvakumaran, N.S.P. Bhuvanesh, R. Karvembu, Self-assembled Cu(II) and Ni(II)
′
′
′
metallamacrocycles formed from 3,3,3 ,3 -tetrabenzyl-1,1 -aroylbis(thiourea)
ligands: DNA and protein binding studies, and cytotoxicity of trinuclear complexes,
Dalton Trans. 43 (2014) 16395–16410.
[60] B. Lal, S. Akhter, A.A. Altaf, A. Badshah, R.A. Hussain, H. Li, Synthesis, crystal
structure, spectral and electrochemical characterization, DNA binding and
antioxidant studies of 1-(2-florobenzoyl)-3-(2-chloro, 4-ferrocenylphenyl)
thiourea, Int. J. Electrochem. Sci. 11 (2016) 1632–1639.
[61] S. Hussain, I. Din, A. Raheel, S. Hussain, M.N. Tahir, I. Hussain, New bioactive Cu
(I) thiourea derivatives with triphenylphosphine; synthesis, structure and
molecular docking studies, J. Coord. Chem. 73 (2020) 1191–1207.
[
[
[62] M. Dehkhodaei, M. Sahihi, H.A. Rudbari, F. Momenbeik, DNA and HSA interaction
of Vanadium (IV), Copper (II), and Zinc (II) complexes derived from an asymmetric
bidentate Schiff-base ligand: multi spectroscopic, viscosity measurements,
1
3

N. Arshad et al.
molecular docking, and ONIOM studies, J. Biol. Inorg. Chem. 23 (2) (2018)
Bioorganic Chemistry 109 (2021) 104707
[64] S. Jahani, M. Khorasani-Motlagh, M. Noroozifar, DNA interaction of europium (III)
′
1
81–192.
complex containing 2, 2 -bipyridine and its antimicrobial activity, J. Biomol.
[
63] N. Shahabadi, S. Mohammadi, Synthesis characterization and DNA interaction
studies of a new Zn (II) complex containing different dinitrogen aromatic ligands,
Bioinorg. Chem. Appl. 2012 (2012).
Struct. Dyn. 34 (3) (2016) 612–624.
[65] N.K. Janjua, A. Shaheen, A. Yaqub, F. Perveen, S. Sabahat, M. Mumtaz, C. Jacob, L.
A. Ba, H.A. Mohammed, Flavonoid–DNA binding studies and thermodynamic
parameters, Spectrochim. Acta A Mol. Biomol. Spectrosc. 79 (5) (2011)
1
600–1604.
1
4