Multicomponent and 1,3-dipolar cycloaddition synthesis of triazole- and isoxazole-acridinedione/xanthenedione heterocyclic hybrids: Cytotoxic effects on human cancer cells

Article abstract of DOI:10.1016/j.molstruc.2020.128325

A new series of diverse 1,2,3-triazole-acridinedione/xanthenedione and 1,2-isoxazole-acridinedione/xanthenedione heterocyclic hybrids have been synthesized via 1,3-dipolar coupling reaction of N/O-substituted-acridinedione-alkyne or O-substituted-xanthene

Full text of DOI:10.1016/j.molstruc.2020.128325

Journal Pre-proof
Multicomponent and 1,3-dipolar cycloaddition synthesis of triazole- and isoxazole-
acridinedione/xanthenedione heterocyclic hybrids: Cytotoxic effects on human cancer
cells
Abdelkader Naouri, Amar Djemoui, Mouhamad Ridha Ouahrani, Mokhtar Boualem
Lahrech, Najet Lemouari, Djenisa H.A. Rocha, Hélio Albuquerque, Ricardo F.
Mendes, Filipe A.Almeida Paz, Luisa A. Helguero, Khaldoun Bachari, Oualid Talhi,
Artur M.S. Silva
PII:
S0022-2860(20)30650-5
DOI:
https://doi.org/10.1016/j.molstruc.2020.128325
MOLSTR 128325
Reference:
To appear in: Journal of Molecular Structure
Received Date: 11 January 2020
Revised Date: 21 April 2020
Accepted Date: 23 April 2020
Please cite this article as: A. Naouri, A. Djemoui, M.R. Ouahrani, M.B. Lahrech, N. Lemouari, D.H.A.
Rocha, Hé. Albuquerque, R.F. Mendes, F.A.A. Paz, L.A. Helguero, K. Bachari, O. Talhi, A.M.S. Silva,
Multicomponent and 1,3-dipolar cycloaddition synthesis of triazole- and isoxazole-acridinedione/
xanthenedione heterocyclic hybrids: Cytotoxic effects on human cancer cells, Journal of Molecular
Structure (2020), doi: https://doi.org/10.1016/j.molstruc.2020.128325.
This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition
of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of
record. This version will undergo additional copyediting, typesetting and review before it is published
in its final form, but we are providing this version to give early visibility of the article. Please note that,
during the production process, errors may be discovered which could affect the content, and all legal
disclaimers that apply to the journal pertain.
© 2020 Published by Elsevier B.V.

Credit author statement:
Naouri Abdelkader conceptualized the work and wrote the original
draft preparation.
Amar Djemoui performed the synthetic experimental work and co-
wrote the manuscript.
Ouahrani Mohammad Ridha and Lahrech Moukhtar Boualem
participated in the synthetic experimental work (purification, isolation
and recrystallisation).
Najet Lemouari and Hélio Albuquerque performed 2D-NMR
analysis and interpretations
Djenisa H. A. Rocha and Luisa A. Helguero performed the
anticancer biological assays
Ricardo F. Mendes and Filipe A. Almeida Paz were responsible for
all X-ray diffraction work.
Khaldoun Bachari, Oualid Talhi and Artur M. S. Silva co-
conceptualized the work and co-wrote the manuscript.

Multicomponent and 1,3-dipolar cycloaddition synthesis of triazole- and
isoxazole-acridinedione/xanthenedione heterocyclic hybrids: cytotoxic effects on
human cancer cells
Abdelkader Naouri,a,b,c,* Amar Djemoui,a,b Mouhamad Ridha Ouahrani,d Mokhtar Boualem Lahrech,b
Najet Lemouari,c Djenisa H. A. Rocha,e,f,g Hélio Albuquerque,e Ricardo F. Mendes,f Filipe A. Almeida
Paz,f Luisa A. Helguero,g Khaldoun Bachari,c Oualid Talhi,c,e, and Artur M. S. Silvae,*
aDepartment of Chemistry, Faculty of Exact Sciences and Informatics ZIANE Achour University, Djelfa, Algeria. abdelkader.naouri@crapc.dz
bLaboratory of Organic Chemistry and Natural Substance, Faculty of Exact Sciences and informatics, ZIANE Achour University, Djelfa, Algeria.
cCentre de Recherche Scientifique et Technique en Analyses Physico-Chimiques, CRAPC, BP384, Bou-Ismail, 42004, Tipaza, Algeria. oualid.talhi@crapc.dz
dDepartment of Chemistry, Faculty of Exact Sciences, Echahid Hamma Lakhdar University of El Oued , Eloued, Algeria.
eQOPNA and LAQV-REQUIMTE, Department of Chemistry, University of Aveiro, 3810-193, Aveiro, Portugal. oualid.talhi@ua.pt, artur.silva@ua.pt
fCICECO-Aveiro Institute of Material, University of Aveiro, 3810-193, Aveiro, Portugal.
gInstitute of Biomedicine (iBiMED), Department of Medical Sciences, University of Aveiro, 3810-193, Aveiro, Portugal.
ABSTRACT
A new series of diverse 1,2,3-triazole-acridinedione/xanthenedione and 1,2-isoxazole-acridinedione/xanthenedione heterocyclic hybrids have been
synthesized via 1,3-dipolar coupling reaction of N/O-substituted-acridinedione-alkyne or O-substituted-xanthenedione-alkyne substrates with
various aromatic azides or oximes. In all cases, the cycloaddition is totally regioselective. The chemical structures of the synthesized compounds
are determined using 2D NMR and are further confirmed by single-crystal X-ray diffraction analysis. Preliminary in vitro cytotoxic assays on two
human breast cancer cell lines (MDA-MB-231, T47-D) and one prostate cancer cell line (PC3) are performed on some selected compounds. The
most active O-1,2,3-triazole-xanthenedione hybrid displays the best cytotoxicity effects with IC50 ≤ 20 µM in breast cancer and IC50 = 10 µM in
prostate cancer cell lines.
Keywords: Click chemistry, Triazole, Isoxazole, Acridinedione, Xanthenedione, Anticancer
———
Corresponding author. e-mail: abdelkader.naouri@crapc.dz, oualid.talhi@ua.pt, artur.silva@ua.pt

2
1. Introduction
The modern trend in organic chemistry is actually the
synthesis of combined organic molecules, especially
heterocycles, as potentially useful structural motifs for drugs
and pharmaceutical industry. Due to the growing need for
potent bioactive compounds for public health, researches are
mostly oriented towards developing newer, more effective
and less toxic molecules showing specific and selective
biological mode of actions and improved properties. In this
regard, a number of new compounds have been designed
bearing various biological activities by the combination of at
least two or more pharmacophores in "one chemical
structure". The use of green chemistry towards the
elaboration of bioorganic-nanomaterial hybrids have
recently evidenced impactful biomedical applications.1
Incorporating five- and six-membered nitrogen and oxygen
containing heterocycles to generate the so-called “hybrid
molecule” has been proved to be effective in terms of
enhanced activity, when compared to that of individual
pharmacophores.1
Figure 1. Examples of interesting structures bearing of
triazole, isoxazole, acridine and xanthene motifs.
In light of the biological and technological importance of
the aforementioned nitrogen and oxygen containing five-
and six-membered heterocycles, it is very interesting to
develop new structures incorporating acridine and xanthene
with 1,2,3-triazole and 1,2-isoxazole motifs in hybrid
molecules. In this context, we describe efficient syntheses of
a novel series of 1,4-disubstituted 1,2,3-triazoles and 3,5-
disubstituted 1,2-isoxazoles by regioselective reaction of
arylazides and aryloximes with the N-/O-acridinedione and
O-xanthenedione derived terminal alkynes via click
chemistry. Preliminary anticancer screening of these hybrid
heterocycles performed on two human breast cancer (MDA-
MB-231, T47-D) and one prostate cancer cell lines (PC3)
shows promising effects
Triazoles and isoxazoles are important classes of nitrogen
and oxygen containing five-membered heterocycles, being
useful in drug design and known as privileged structures.2
Triazoles are mostly applied in medicinal,3 pharmaceutical,4
biological5 and material sciences.6 This heterocyclic motif is
found as a fingerprint of some known drugs such as
fluconazole7 (Figure 1). Triazoles and isoxazoles have a
wide range of biological properties such as anticancer,8 anti-
HIV,9 antibacterial,10 antimalarial11 and anti-tubercular12
activities. Particularly, triazoles are stable to hydrolysis,
oxidation and relatively resistant to metabolic degradation.13
On the other side, isoxazoles are found as the basic structure
of remarkable drugs, for example, the antibiotic
sulfamethoxazole14 (Figure 1).
2. Results and discussion
2.1. Chemistry and spectral analysis
We first describe the synthesis of N-1,2,3-triazole-
dioxodecahydroacridine
11a-e
and
N-1,2-isoxazole-
dioxodecahydroacridine 12a-e heterocyclic hybrids via Click
reaction. The procedure begins by the preparation of 1,8-dioxo-
decahydroacridine derivatives via a one-pot three-component
condensation of aromatic aldehydes 1a-e, 1,3-cyclohexanedione
2 and ammonium acetate 3 in ethanol catalyzed by triethylamine
(TEA) to give the desired intermediates 4a-e in excellent yields
(86-96%) (Scheme 1).26 Then the N-alkylation of 1,8-
dioxodecahydroacridines 4a-e with propargyl bromide 5 in
presence of K2CO3 as a base and DMF as solvent at room
temperature were carried out. The reaction normally takes place
in 3 hours to afford the corresponding N-propargyl-substituted
1,8-dioxodecahydroacridines 6a-e obtained in moderate yields
(34-55%, Scheme 1). In the following step, arylazides 8a,b were
prepared by reacting aniline derivatives 7a,b with sodium nitrite
and sodium azide in acidic medium at room temperature. The
reaction of aldehydes 9a,b with hydroxylamine hydrochloride in
refluxing water under basic (NaHCO3) conditions provides the
corresponding aryloximes 10a,b (Scheme 2).
A variety of synthetic routes have been reported for the
synthesis of triazoles and isoxazoles, being one of the most
popular the [3+2] cycloaddition of alkynes with azides or
nitrile oxides, respectively.15 In parallel, we found that the
multicomponent reaction (MCR) approach has been widely
used for the production of complex and highly diverse
nitrogen and oxygen containing six-membered heterocycles
such as acridine and xanthene. MCRs are experimentally
performed in shorter time and in a single step allowing
excellent yields and highly pure compounds. By using easily
accessible starting materials, outstanding acridine templates
containing
1,4-dihydropyridines
(1,4-DHPs)
and
xanthenediones with a pyrane ring have been accessed via
MCRs without the isolation of reaction intermediates.15
These structures have also shown a wide range of
pharmacological and biological activities.16 DHPs were
firstly developed as cardiovascular agents17, but they have
also found applications for other medicinal treatments, for
instance, nifedipine which is employed in the treatment of
migraine, hypertrophic cardiomyopathy and Raynaud’s
phenomenon18 (Figure 1). Among the various subclasses of
pyrans, 9-arylxanthenes are active oxygenated heterocycles,
which are important drug intermediates. They are known as,
anti-inflammatory,19 antiviral,20 antibacterial21 agents and
thoroughly used in photodynamic therapy to destroy tumor
cells.22 Xanthene derivatives are widely utilized in laser
technology because of their interesting spectroscopic
properties,23 in fluorescent materials for the visualization of
biomolecules24 and especially as dyes like the popular
Scheme 1. Synthesis of 1,8-dioxodecahydroacridines 4a-e
and their N-propargylated derivatives 6a-e.
rhodamine 6G25
(Figure 1).

Scheme 2. Preparation of arylazides 8a,b and aryloximes
10a,b
.
The last step settles in a simple 1,3-dipolar cycloaddition
reactions between N-propargyl-substituted-1,8-
dioxodecahydroacridines 6a-d with aromatic azides 8a,b, in the
presence of CuSO4.5H2O/ascorbic acid as catalyst in a 2:1
mixture of dichloromethane:H2O at room temperature for 12 h to
produce 1,2,3-triazole-dioxodecahydroacridine hybrids 11a-e in
good to excellent yield (70-86%) (Scheme 3). On the other hand,
subjecting N-propargyl-substituted 1,8-dioxodecahydroacridines
6a-d to [3+2] cycloaddition reaction with various aryloximes
10a,b in the presence of chloramine-T, CuSO4 and Cu powder
furnished the desired 1,2-isoxazole-dioxodecahydroacridine
hybrids 12a-e (65-80%) (Scheme 3).
1
13
The combination of H and C NMR spectroscopy allows a
clear determination of the regioselective formation of a
unique 1,4-disubstituted 1,2,3-triazole ring in the 1,2,3-
triazole-N-acridinedione and 1,2-isoxazole-N-acridinedione
hybrids. The characteristic signal of the triazolic H-5” of
compound 11a appears as a singlet at δH 8.75 ppm, while
that of isoxazole 12a showed the singlet H-4’’ around δH
6.89 ppm. Aliphatic signals at δH 5.18, 5.29 ppm are
assigned to the methylene H-1’ linking the heterocyclic
moieties (triazole or isoxazole) to the acridine bulk. 13C
NMR of 11a displayed the triazole skeleton with C-5” at δC
121.8 and the quaternary C-4” at δC 145.2 ppm based on its
HMBC correlations established with H-1’ and H-5” (Figure
2). For the isoxazole 12a, one protonated carbon C-4” (δC
101.0 ppm) and two different quaternary carbons C-3” and
C-5” are distinguished, being respectively attributed to δC
162.10, 169.4 ppm and confirmed through HMBC
correlations entertained between C-3”/H-4” and C-5”/H-
1’/H-4” (Figure 2). Other important HMBC cross-peak
correlations have been studied, which greatly help us to establish
the symmetrical structure of acridine, especially those observed
between H-9’ with its neighbouring quaternary carbons C8a/9a
Scheme
dioxodecahydroacridines
dioxodecahydroacridines 12a-e via click reaction.
3.
Synthesis
11a-e
of
and
N-1,2,3-triazole-
N-1,2-isoxazole-
Good quality single-crystals have only been successfully
obtained for the isoxazole-acridine template 12a, being isolated
from a 1:1 mixture of hexane:dichloromethane by slow
evaporation at ca. 6 °C. The crystal structure was determined in
the centrosymmetric triclinic Pī space group. The asymmetric
unit is composed of a whole molecular unit of 12a plus partially-
occupied water and dichloromethane solvent molecules (Figure
3). Remarkably, though the molecule is rich in atoms capable of
being engaged in strong hydrogen bonds and acceptors, there is a
lack of donating moieties (besides the disordered and partially-
occupied water molecules of crystallization). This leads to a
general weak supramolecular network which also accounts for
the various structural disorder features found in the crystal
structure (ESI for additional technical details).
(
δC 115.2
carbonyl C-1 (δC 195.7-195.8 ppm) of both hybrid structures
11a and 12a (Figure 2, ESI for NMR spectra)
-116.0 ppm), C-4a/10a (δC 153.9-154.2 ppm) and the
Further synthetic work was focused on the elaboration of 1,4-
disubstituted
and
1,2,3-triazole-O-acridinedione/O-xanthenedione
1,2-isoxazole-O-acridinedione/O-
.
3,5-disubstituted
xanthenedione hybrid compounds, and this time, we start with
the alkylation of commercially available hydroxybenzaldehydes
13a-d using propargyl bromide in the presence of K2CO3 to give
the corresponding propargyloxybenzaldehydes 14a-d, bearing an
alkynyl group required for click chemistry (Scheme 4).
Compounds 14a-d are then subjected to click chemistry, as
previously reported by employing arylazides 8a,b in the presence
of CuSO4·5H2O and sodium ascorbate as the catalytic system in
dichloromethane:H2O (2:1, v/v) at room temperature for 12 h.
This provides 1,4-disubstituted 1,2,3-triazolealdehydes 15a-e in
good yields (55-88%).27

4
Figure 3. Schematic representation of the molecular units present
in the crystal structure of compound 12a·0.8CH2Cl2·0.4H2O.
Non-hydrogen atoms are represented as thermal ellipsoids drawn
at the 50% probability level (except for those which were refined
isotropically – see ESI for additional technical details), and
hydrogen atoms as small spheres with arbitrary radii.
Scheme 4. Synthesis of propargyloxybenzaldehydes 14a-c.
Figure 2. Main HMBC correlation observed for N-1,2,3-triazole-
In the same fashion as mentioned above, we prepare 1,2-
isoxazolealdehydes 16a-e by click reaction of aryloximes 10a,b
with propargyloxyaldehydes 14a-d in the presence of
chloramine-T, CuSO4 and Cu powder as a catalyst, being the
desired 1,2-isoxazolealdehydes 16a-e also obtained in good
yields (72-92%). The ending step of our synthetic strategy aims
at the production of 1,2,3-triazole-O-acridinediones 17a-e and
1,2-isoxazole-O-acridinediones 19a-e hybrids by a Hantzsch
reaction. We therefore performed the one-pot three-component
reaction between two molecules of 1,3-cyclohexanedione, the
corresponding triazole-aldehydes 15a-e or isoxazole-aldehydes
16a-e and ammonium acetate using triethylamine as a catalyst in
ethanol.
dioxodecahydroacridine
dioxodecahydro-acridine 12a.
11a
and
N-1,2-isoxazole-
The target heterocyclic hybrids 17a-e and 19a-e were isolated in
good to excellent yields (55-90%) (Scheme 5).26 In addition, we
have investigated another MCR route for the synthesis of 1,2,3-
triazole-O-xanthenediones
18a-e
and
1,2-isoxazole-O-
xanthenediones 20a-e hybrids by using two molecules of 1,3-
cyclohexanedione and the corresponding triazole-aldehydes 15a-
e or isoxazole-aldehydes 16a-e with triethylamine as catalyst in
acetic acid to afford the desired compounds 18a-e and 20a-e in
very good yields (67-91%)28 (Scheme 5).

Scheme 5. Synthesis of
acridinediones 19a-e and
O
-1,2,3-triazole-acridinediones 17a-e
, O-1,2,3-triazole-xanthenediones 18a-e, O-1,2-isoxazole-
O-1,2-isoxazole-xanthenediones 20a-e
.
which help to characterize the hybrid structures 17-20
(Figure 4, ESI for NMR spectra).
The examination of the NMR spectra of our novel heterocyclic
hybrids 1,2,3-triazole-O-acridinediones/O-xanthenediones 17a-e,
18a-e and 1,2-isoxazole-O-acridinediones/O-xanthenediones
19a-e, 20a-e suggests that most of the pure products are obtained
in a regioselective manner affording 1,4-disubstituted 1,2,3-
triazoles and 3,5-disubstituted isoxazoles linked to acridinediones
To support our regioselective preparation of the reported
heterocyclic hybrids, we crystallised selected compounds to be
investigated by single-crystal X-ray diffraction. We isolated good
crystals of the O-isoxazole-acridine hybrid 19b from an ethanolic
solution by slow evaporation at ca. 6 °C. The crystal structure
was solved and refined in the centrosymmetric monoclinic P21/n
space group, with the asymmetric unit being composed of a
whole molecular unit of 19b and a partially-occupied water
molecule of crystallisation (refined as 75%, Figure 5). Though
the molecular unit is rich acceptor atoms capable of engaging in
strong and directional hydrogen bonding interactions, there is
only one donor group capable of such (besides the partially-
1
or xanthenediones. From the H NMR and 2D-HSQC spectra of
these compounds, we could distinguish two characteristic groups
9-CH (δH 4.54-5.15 ppm, δC 30.4-32.0 ppm) and 1”-CH2 (δH
5.14-5.26 ppm, δC 60.5-62.0 ppm), being considered as the
linking bridges between the acridine or xanthene skeleton and the
triazole or isoxazole heterocycles. The triazole and isoxazole
protons H-5”’ and H-4”’ appear as a singlet at δH 8.89-8.90 and
7.12-7.16 ppm, respectively. The cyclohexanone parts can be
characterized through their aliphatic protons at δH 1.78-1.90 ppm
for H-3/6, 2.11-2.28 ppm for H-2/7 and 2.43-2.53 ppm for H-4/5
(case of compound 17a).
occupied water molecule of crystallization): the –NH moiety of
the dioxodecahydroacridine. This moiety is indeed engaged in an
unique bifurcated N–H···O interaction with the oxygen atoms of
a neighboring molecular unit [dN···O ranging from 2.892(4) to
3.156(3) Å; <(NHO) interaction angles in the 115(3)-178(3)º
range], forming a C11(10) graph set motif.29
From the 13C NMR and 2D-HSQC spectra, all the
protonated carbon have been successfully assigned, mainly
those of the cyclohexanone structure δC 21.3 ppm for C-3/6,
26.8 ppm for C-4/5 and 37.2 for C-2/7 (case of compound
17a). However, it was possible to differentiate between
these aliphatic carbons only by interpreting the 2D-HMBC
spectrum of compound 17a, where we find 10-N
H (δH 9.37
ppm) establishing remarkable J3 HMBC correlation with
H/C
the aliphatic carbon C-4/5. Other interesting HMBC
connectivities have been observed for the 10-N
H with the
quaternary carbons of the cyclic acridine skeleton, where we
could attribute C8a/9a to δC 113.1 ppm via J3H/C correlation of
10-NH and C-4a/10a to δC 151.5 ppm via J2 with this same
H/C
proton. It was also possible to discover a rare J4
H/C
connection of 10-N
H with the carbonyl C-1 (δC 195.2 ppm),

6
4""
4""
have no considerable effect, while 10% of inhibitory could
be observed for compounds 11d 17c 17d 18e and 20b. In
the other side, compounds 17d, 20a 20b and 20e are less
active on the breast cancer (T47-D and MDA-MB-231). The
most active compounds 11e 12a 18a exhibited total or
near-total cytotoxicity at 100 µM concentrations, but no
effects were observed at concentrations lower than 10 µM.
Therefore, the 10-100 µM concentration range was
expanded to cover values every 0.25 unit to allow
calculation of the IC50 values as shown in Table 1.
,
,
,
,
H
H
3"'
2""
N
2""
1"'
N
N
3"' N
1"'O
H
H
,
,
-d
H
1"
1"
O
4'
H
H
O
4'
H
OCH3
2'
2'
H
H
O
O
O
O
1
H
H
H
8a
10a
H
9
8a
10a
1
9
9a
4a
9a
4a
6
4
6
4
N
H
H
N
H
H
17a
19b
Figure 4. Main HMBC correlations of N-1,2,3-triazole-dioxo-
decahydroacridine 17a and N-1,2-isoxazole-dioxodecahydro-
acridine 19b.
Figure 6. Representative graphic of in vitro cytotoxicity
assessment of compounds 11a-d, 17c-d, 18e, 19b, 19e, 20a-b
and 20e against breast cancer (T47-D and MDA-MB-231) and
prostate cancer (PC3) cell lines at concentration of 100 µM.
The results showed that in metastatic cancer cell lines
(MDA-MB-231) compounds 12a, 18b and 18c displayed
IC50 value ≤ 20 µM. Compounds 18b-d exhibited the
highest cytotoxic effect against the non-metastatic cell line
(T47-D) with IC50 value ≤ 20 µM (Table 1). The in vitro
antiproliferative
activity
of
N-1,2,3-triazole-
dioxodecahydroacridines 11a
-e in both cell lines tested are
weak, nevertheless their inhibitory effects are more
pronounced in the non-metastatic cells (T47-D) than in the
Figure 5. Schematic representation of the molecular unit present
in the crystal structure of compound 19b·0.75H2O. Non-
hydrogen atoms are represented as thermal ellipsoids drawn at
the 70% probability level (except for those associated with the
disordered methyl group – see the ESI for additional technical
details), and hydrogen atoms as small spheres with arbitrary
radii.
metastatic cells (MDA-MB-231 and PC3) (Figure 6). The
1,2-isoxazole-dioxodecahydroacridine 12a showed similar
cytotoxic effect in T47-D cells comparing to compound 11e
N-
.
However, compound 12a exhibited around 6-fold more
potency in MDA-MB-231 cell than compound 11e (Table
1). These results suggested that the introduction of 1,2-
isoxazole molecule at the acridine scaffold may lead to an
increased cytotoxicity in metastatic cancer cells. Compounds
12b-e should be evaluated to confirm these results, however
due to some difficulties of solubility in DMSO, the
experiments were not performed.
2.2. In vitro evaluation of cytotoxic effects
The
cytotoxicity
of
N-1,2,3-triazole-
dioxodecahydroacridines
dioxodecahydroacridines
11a
12a
-
e
,
,
N
O
-1,2-isoxazole-
-1,2,3-triazole-
acridinediones 17c
-1,2-isoxazole-acridinediones 19b
isoxazole-xanthenediones 20a-b 20e have been studied in
-
d
,
O
-1,2,3-triazole-xanthenediones 18a-
The preliminary evaluation of in vitro antiproliferative activity
e,
O
,
19e and -1,2-
O
of compounds 17c
that he most active hybrids are
xanthenediones 18a-d Among them, compound 18c without
-d
,
18a-e
,
19b
,
19e
,
20a-b and 20e showed
,
t
O-1,2,3-triazole-
two human breast cancer cell lines (MDA-MB-231, T47-D)
and one prostate cancer cell line (PC3) after 72 h of
exposure using the prestoblue method.30 Compounds have
been selected upon their optimized solubility in DMSO,
hence we started the biological screening by using a 10 nM
to 100 µM concentrations curve with a dilution factor of 10
(doxorubicin is employed as a positive control). Most of the
selected compounds displayed less than 60% of inhibitory
effects on the tested cancer cell lines at 100 µM (Figure 6).
In prostate cancer cells (PC3), compounds 11c, 19e and 20a
.
any substituents in the aromatic rings displayed the most
effective cytotoxicity on the tested cancer cell lines with IC50
values less than 20 µM in case of breast cancer (MDA-MB-231
and T47-D) and have recorded the highest cytotoxic effect in
PC3 cells, with IC50 value of 10 µM (Table 1). These results
suggest that the ortho position of the 1,2,3-triazole motif at the
xanthenedione scaffold favours the cytotoxicity in PC3
cancer cells when comparing to compound 18a
, 18b and
18d of the same group. In case of -1,2,3-triazole-
O

MCF-7 [R = Cl (1.20±0.13)
R =3,4,5-OCH3 (0.23±0.02)
R = CF3 (0.17±0.01)]
acridinedione 17 and
O-1,2-isoxazole-acridinedione 19, no
significant inhibitory effects were observed in the tested
cancer cells at concentrations below 100 µM. At 100 µM
compound 19b is clearly affecting all cancer cell lines
compared to 17d (Figure 6), which may inform that the
conjugation and position of the isoxazole moiety at the
acridinedione scaffold is in favour of an increased
cytotoxicity.
HT-29 [R = Cl (1.10±0.09)
R =3,4,5-OCH3 (0.11±0.03)
R = CF3 (2.54±0.19)]
A-549 [R = Cl (0.28±0.03)
R =3,4,5-OCH3 (1.90±0.96)
R = CF3 (1.45±0.18)]
A-375 [R = Cl (0.39±0.02)
R =3,4,5-OCH3 (1.23±0.11)
R = CF3 (1.67±0.12)]
Table 1. In vitro cytotoxicity assessment of compounds 11e, 12a,
18a-d against breast cancer (T47-D and MDA-MB-231) and
prostate cancer (PC3) cell lines.
HCT-116 (23.11±1.03)
Compound
T47-D
MDA-MB-
231
PC3
IC50 µM(a)
IC50 µM(a)
IC50 µM(a)
65.35±3.18
10.78±0.85
49.45±1.37
17.69±1.18
20.88±0.20
45.31±0.87
1.51±0.97
11e
12a
18a
18b
18c
18d
38.05±4.10
49.02±1.03
26.70±0.44
18.40±3.89
14.50±1.59
19.40±1.67
58.19±3.57
38.65±4.50
25.86±0.56
64.58±2.11
10.20±0.22
66.84±1.09
0.73±0.14
ER-positive MCF-7 (12.4 ±
0.54 μM)
ER–negative MDA-MB-231
cells (25.4 ± 0.40 μM)
Doxorubicin 0.13±0.003
[a] IC50 determined by the prestoBlue method after 72h of
incubation. Each value is the mean (IC50±SD) of two
independent experiments performed in quadruplicate
The cytotoxic activity of reported hybrid compounds sharing
acridine/xanthene have resulted in promising antiproliferative
activity against a panel of tested cancer cells (Table 2).31-34 The
hybrid acridine-1,2,4-triazole compunds with different
substitutions at the para position of the phenyl ring exhibited the
most potent anticancer activity against four cancer cell lines
(MCF-7, HT-29, A-549, A-375) with IC50 values ≤ 5 μΜ.32
Acridine-thiophene hybrids shows selectivity toward HCT-116
cells, whereas no cytotoxicity was observed for other cell lines
tested (HeLa, MCF-7, K562, HL-60, HaCat, PBMC).33 Synthetic
xanthene-pyrrolidine was reported to possess comparable
behavior to Tamoxifen on ER-positive MCF-7 and ER–negative
MDA-MB-231 cells (Table 2).34 Our data screening of cytotoxic
3. Conclusion
In summary, novel series of acridinediones and
xanthenediones containing triazole and isoxazole moieties
with varied aromatic substitution, all in one hybrid structure,
have been synthesized in appreciable yields. The spectral
NMR data and single-crystal X-ray have been well exploited
to finely characterize the molecular structures with various
structural patterns. Different synthetic methodologies have
been combined from a one-pot multi-component reaction to
1,3-dipolar cycloaddition and click chemistry in order to
achieve a high structural complexity of novel heterocyclic
hybrids. The preliminary anticancer screenings on human
cancer cell lines demonstrate that the
O-1,2,3-triazole-
effect
of
1,2,3-triazole
and
1,2-oxazole-
xanthenediones hybrids show the best cytotoxic effects.
Moreover, N,O-isoxazole moiety coupled to the
acridinedione scaffold could lead to an increased
cytotoxicity. Further antiproliferative studies are required to
provide more insights about the combination of heterocyclic
pharmacophores in a hybrid molecule and their structure-
activity-relationship in cancer therapy.
acridinediones/xanthenediones heterocyclic hybrids provide
essential SAR information and experimental structural models
for anticancer drug design.
Table 2. Reported studies of in vitro cytotoxicity assessment of
some hybrid compounds against a panel of cancer cell lines.
Organic hybrid
structures
Cancer cell lines tested
(IC50±SD μM)
7 (16.24± 0.54)
A2780 (10.43±1.46)
HeLa (22.86±1.45)
HepG2 (15.07±1.63)
DU145 (13.73±1.26)
A549(9.01±0.24)
PC3 (5.23±1.37)
LNCAP (18.54±2.11)
HUVEC (49.19±2.3)
4. Experimental section
MCF
‐
General remarks: Melting points were measured using
Kofler bench method. All reactions were followed by TLC
(E. Merck Kieselgel 60 F-254), with detection by UV light
1
at 254 nm. H and 13C NMR spectra were recorded on a
Bruker AC 400 MHZ FTNMR spectrometer using DMSO-
d6 as solvent. Chemical shifts (δ) were reported in parts per
million (ppm) relative to tetramethylsilane TMS (δ = 0 ppm)
used as an internal reference and coupling constants (J) were
given in Hz. The following multiplicity abbreviations were
used: s, singlet; d, doublet; t, triplet; q, quadruplet; m,
multiplet. High-resolution mass spectra (ESI+-HRMS) were
measured with a micrOTOF-Q 98 spectrometer. All
chemicals and solvents were purchased from commercial
sources and used as received.
General procedure for the synthesis of 1,8-
dioxodecahydroacridines (4a-e, 17a-e and 19a-e).
A

8
mixture of aldehyde 1a-e (or 15a-e
cyclohexane-dione (2 mmol, 0.224 mg), ammonium
acetate (3 mmol, 0.231 mg), triethylamine (2 mmol, 0.202
/
16a-e) (1 mmol), 1,3-
mmol, 0.455 mg) was gradually added over 3 minutes
followed by CuSO4.5H2O (1 mmol, 0.250 mg) and copper
powder (2 mmol, 0.128 mg). To the aforementioned
solution, the appropriate 1,8-dioxodecahydroacridine
2
3
mg) in ethanol (20 mL) was placed in a 50 mL flask and
refluxed under continuous stirring for the appropriate time
as monitored by thin-layer chromatography TLC. After
completion of the reaction, the solvent was evaporated to
give the crude product, which was purified by
recrystallization from EtOH to provide the pure product 4a-e
derived alkyne 6a-e (or propargyloxybenzaldehydes 14a-e
)
(1 mmol) was added and the reaction mixture was stirred for
12 hours at room temperature. Water (10 mL) was added to
the reaction mixture and directly extracted with
dichloromethane (3 × 15 mL). The organic layer was dried
over anhydrous sodium sulfate, filtered, concentrated and
then recrystallized from ethanol to give the required
isoxazoles 12a-e (or 16a-e).
(17a-e / 19a-e) without further purification.
General procedure for the synthesis of
1,8-dioxodecahydroacridines
N-propargylated
and
propargyloxybenzaldehydes (6a-e and 14a-e). A mixture
of 1,8-dioxodecahydroacridine 4a-e (or
hydroxybenzaldehyde 13a-e derivatives (2 mmol),
propargyl bromide (2.2 mmol, 0.262 mg) and K2CO3 (4
mmol, 0.552 mg) in -dimethylformamide (15 mL) was
stirred at room temperature for 3 hours. Water (15 mL) was
added to the reaction mixture to be extracted with
dichloromethane (3 x 15 mL). The organic layer was dried
over anhydrous sodium sulphate, filtered and concentrated.
The final residue was recrystallized from ethanol to afford
General procedure for the synthesis of 1,8-dioxo-
octahydroxanthenes 18a-e and 20a-e. A mixture of
)
aldehyde 15a-e (or 16a-e
)
(0.3 mmol), 1,3-
5
cyclohexanedione (0.6 mmol, 0.067 mg) and triethylamine
2
N
,
N
(0.6 mmol, 0.061 mg) as catalyst in 10 mL of acetic acid
was allowed to stir at reflux for the appropriate time. After
completion of the reaction under TLC monitoring, the
solvent was evaporated under vacuum and the solid product
obtained was washed with ethanol and then water. The
products were purified by recrystallization from ethanol to
the desired compounds 6a-e (or 14a-e
).
give the desired compounds 18a-e
(20a-e).
General synthetic procedure for the aromatic azides
8a,b. The aniline substrate 7a,b (10 mmol) was dissolved in
hydrochloric acid (15 mL) in a round-bottom flask and
9-(4-Methoxyphenyl)-10-[(1-phenyl-1H-1,2,3-triazol-4-
yl)methyl]-3,4,6,7,9,10-hexahydroacridine-1,8(2 ,5 )-
H H
dione (11a): C29H28N4O3; MW = 480.56 g/mol; 0.344 g
1
cooled to 0
⁰C in an ice bath. Ice cooled sodium nitrite
(69.5%); yellow solid; mp 258-260 °C. H NMR (400 MHz,
NaNO2 (12 mmol, 0.830 mg) solution was added into the
aniline-acid solution and stirred for 10 min. NaN3 (12 mmol,
0.780 mg) was then added under stirring for 1 hour. Finally,
the solution was extracted with ethyl acetate (3 x 15 mL)
and the organic layers were combined and dried over
Na2SO4. After solvent removal under reduced pressure, the
azidobenzene derivatives 8a,b are afforded.
DMSO-d6) δ : 1.85-1.98 (2m, 4H, H-3/6, eq:ax), 2.21–2.28
(m, 4H, H-2/7), 2.64-3.08 (2m, 4H, H-4/5, eq:ax), 3.57 (s,
3H, 4””-OCH3), 5.02 (s, 1H, H-9), 5.18 (s, 2H, H-1’), 6.58
(d,
J = 8.7 Hz, 2H, H-3””/5””), 6.91 (d, J = 8.7 Hz, 2H, H-
2””/6””), 7.40-7.55 (m, 1H, H-4”’), 7.56-7.70 (m, 2H, H-
3”’/5”’), 7.83-7.99 (m, 2H, H-2”’/6”’), 8.75 (s, 1H, H-5”)
13
ppm. C NMR (100 MHz, DMSO-d6) δ : 21.5 (C-3/6), 26.7
(C-4/5), 30.1 (C-9), 36.4 (C-2/7), 41.1 (C-1’), 55.4 (4””-
OCH3), 113.5 (C-3””/5””), 116.0 (C-8a/9a), 120.4 (C-
2”’/6”’), 121.8 (C-5”), 128.7 (C-2””/6””), 129.4 (C-4”’),
130.3 (C-3”’/5”’), 136.9 (C-1”’), 139.1 (C-1””), 145.2 (C-
4’’), 153.9 (C-4a/10a), 157.6 (C-4””), 195.8 (C-1/8, C=O)
ppm. HRMS (ESI+): m/z calcd for [C29H28N4O3+H]+:
481.2161; found: 481.2234.
General procedure for the synthesis of 1,2,3-triazole
derivatives via 1,3-dipolar cycloaddition (11a-e and 15a-
c). Synthetic phenyl azide derivatives 8a,b (1 mmol) and
1,8-dioxodecahydroacridine derived alkyne 6a-e (or
propargyloxybenzaldehydes 14a-e) (1 mmol) were taken in
CH2Cl2 and H2O (10 mL, 1:1) system in a 50 mL round
bottomed flask. CuSO4.5H2O (2.2 mmol, 0.548 mg) and
sodium ascorbate (2.8 mmol, 0.554 mg) were added. The
reaction mixture was stirred at room temperature for 12
hours under TLC monitoring. After completion of the
reaction, the aqueous layer was extracted with CH2Cl2 (3 ×
15 mL). The combined organic phases were dried over
Na2SO4 and concentrated under reduced pressure.
The obtained crude product was recrystallized from ethanol
to afford the pure products 11a-e (or 15a-e).
9-(4-Chlorophenyl)-10-{[3-(4-methoxyphenyl)isoxazol-5-
yl]methyl}-3,4,6,7,9,10-hexahydroacridine-1,8(2H,5H)-
dione (12a): C30H27ClN2O4; MW = 515.01 g/mol; 0.331 g
1
(64.3%); white solid; mp 142-144°C. H NMR (400 MHz,
DMSO-d6) δ : 1.84-1.98 (2m, 4H, H-3/6, eq:ax), 2.18–2.31
(m, 4H, H-2/7), 2.57-2.96 (2m, 4H, H-4/5, eq:ax), 3.83 (s,
3H, 4”’-OCH3), 5.10 (s, 1H, H-9), 5.29 (s, 2H, H-1’), 6.89
(s, 1H, H-4”), 7.03–7.14 (m, 4H, H-3”’/5”’, H-2””/6””),
7.17 (d,
J = 8.5, 2H, H-3””/5””), 7.78 (d, J = 8.8, 1H, H-
General synthetic procedure for the aldoximes (10a,b).
To a methanolic solution of aldehyde 9a,b (10 mmol),
hydroxyl amine hydrochloride (10 mmol, 0.695 mg) was
added followed by sodium acetate (10 mmol, 0.770 mg).
The mixture was stirred at room temperature. After
completion of the reaction as monitored by TLC, water was
added to the reaction mixture and then extracted with ethyl
acetate (3 × 15 mL). The organic layers were combined and
dried over Na2SO4. After solvent removal under reduced
pressure aldoxime derivatives 10a,b are afforded
2”’/6”’) ppm. 13C NMR (100 MHz, DMSO-d6) δ(ppm): 21.4
(C-3/6), 26.3 (C-4/5), 30.6 (C-9), 36.5 (C-2/7), 42.0 (C-1’),
55.8 (4”’-OCH3), 101.0 (C-4”), 115.1 (C-3”’/5”’), 115.2 (C-
8a/9a), 120.9 (C-1”’), 128.2 (C-3””/5””), 128.6 (C-2”’/6”’),
129.5 (C-2””/6””), 130.9 (C-4””), 145.4 (C-1””), 154.2 (C-
4a/10a), 161.2 (C-4”’), 162.10 (C-3”), 169.4 (C-5”), 195.7
(C-1/8, C=O) ppm. HRMS (ESI+): m/z calcd for
[C30H27ClN2O4+H]+: 515.1659; found: 515.1732.
9-{4-[(1-Phenyl-1
3,4,6,7,9,10-hexahydroacridine-1,8(2
C28H26N4O3; MW = 466.54 g/mol; 0.409 g (87.6%); white
solid; mp > 260 °C. H NMR (400 MHz, DMSO-d6) δ 1.78-
1.90 (2m, 4H, H-3/6, eq:ax), 2.11–2.28 (m, 4H, H-2/7),
2.43-2.53 (m, 4H, H-4/5, eq:ax), 4.86 (s, 1H, H-9), 5.14 (s,
H
-1,2,3-triazol-4-yl)methoxy]phenyl}-
,5 )-dione (17a):
H
H
General procedure for the synthesis of 1,2,-isoxazole
derivatives via 1,3-dipolar cycloaddition (12a-e and 16a-
e). To the appropriate aldoximes 10a,b (1 mmol) dissolved
1
in 10 mL of t-BuOH/H2O (1:1), chloramine-T trihydrate (2

2H, H-1”), 6.86 (d,
J
= 8.7, 2H, H-3’/5’), 7.08 (d,
J
= 8.7,
Cell culture: MDA-MB-231 metastatic breast cancer cell
line was grown in DMEM culture medium supplemented
with 10% fetal bovine serum (FBS) (GibcoTM by Life
Technology) and 1% penicillin/streptomycin solution
(PEST; GibcoTM). T47-D non-metastatic luminal breast
cancer and PC3 metastatic prostate cancer cell lines were
grown in RPMI culture medium supplemented with 10%
FBS and 1% PEST. Cell cultures were maintained at 37 ˚C
in a 5% CO2 humidified atmosphere.
2H, H-2’/6’), 7.45-7.53 (m, 1H, H-4”’), 7.55-7.64 (m, 2H,
H-3”’/5”’), 7.86-795 (m, 2H, H-2”’/6”’), 8.89 (s, 1H, H-
5”’), 9.37 (s, 1H, 10-NH) ppm. 13C NMR (100 MHz,
DMSO-d6) δ : 21.3 (C-3/6), 26.8 (C-4/5), 31.6 (C-9), 37.2
(C-2/7), 61.5 (C-1”), 113.1 (C-8a/9a), 114.4 (C-3’), 120.6
(C-2””/6””), 123.3 (C-5”’), 128.9 (C-2’), 129.2 (C-4””),
130.3 (C-3””/5””), 137.0 (C-1””), 140.6 (C-1’), 144.6 (C-
4”’), 151.5 (C-4a/10a), 156.4 (C-4’), 195.2 (C-1/8, C=O)
ppm. HRMS (ESI+): m/z calcd for [C28H26N4O3+H]+:
467.2005; found: 467.2078.
Reagent and tested compounds: Compounds were
dissolved in 100% DMSO to obtain a stock concentration of
100 mM. Thereafter, serial dilutions were prepared in
growth medium. Maximum DMSO concentration applied to
the cells was 0.1% v/v to avoid toxic effects associated to
higher concentrations of this solvent.
9-{4-[(1-(4-Methoxyphenyl)-1H-1,2,3-triazol-4-
yl)methoxy] phenyl}-3,4,5,6,7,9-hexahydro-1
H-xanthene-
1,8(2 )-dione (18b): C29H27N3O5; MW = 497.55 g/mol;
H
1
0.126 g (84.4%); brown solid; mp 178-182 °C. H NMR
(400 MHz, DMSO-d6) δ 1.74–2.02 (2m, 4H, H-3/6, eq:ax),
2.15–2.37 (m, 4H, H-2/7), 2.53–2.74 (m, 4H, H-4/5, eq:ax),
3.83 (s, 3H, 4””-OCH3), 4.54 (s, 1H, H-9), 5.14 (s, 2H, H-
1”), 6.81–6.98 (m, 2H,H-3’), 7.12 (m, 4H, H-2’/3””), 7.69–
7.89 (m, 2H, H-2””), 8.79 (s, 1H, H-5”’) ppm. 13C NMR
(100 MHz, DMSO) δ 20.4 (C-3/6), 26.9 (C-4/5), 30.4 (C-9),
36.9 (C-2/7), 56.0 (4””-OCH3), 61.5 (C-1”), 114.6, (C-3’),
115.3, B (C-3’”), 116.2 (C-8a, C-9a), 122.3 (C-2””), 123.2
(C-5”’), 129.5, (C-2’), 130.5, (C-1””), 137.7 (C-1’), 144.2,
(C-4”’), 156.9 (C-4’), 159.8 (4””-OCH3), 165.1 (C-4a, C-
10a), 196.8 (C-1/8, C=O) ppm. HRMS (ESI+): m/z calcd for
[C29H27N3O5+H]+: 498.1951; found: 498.2023.
Cell viability assay: Serial dilutions ranged from 10nM
to 100 µM are prepared to cover a wide scale for generation
of dose-response curves. In all experiments, the solvent
DMSO (0.1% v/v) alone was used as negative control. Ten
thousand cells/wells were seeded in 96-well plates and
allowed to adhere for 24 h. Cells were then exposed to the
test compounds diluted in culture medium for 48 hours (200
µL), after this time 100 µL of the culture medium was
replaced by test solution for additional 24 h. Thereafter, cell
viability was assessed using 100 µL/well of the rezasurin-
based prestoBlueTM reagent (Life Technologies) according
to the manufacturer´s instructions. Values considered after 3
h of incubation were within the linear range of the reading.
The IC50 values from at least three independent experiments
were calculated using GraphPad Prism software (version
6.00), using the log (inhibitor) vs. response (variable slope-
four parameters) function.
9-(3-Methoxy-4-[(3-phenylisoxazol-5-
yl)methoxy]phenyl)-3,4,6,7,9,10-hexahydroacridine-
1,8(2H,5H)-dione (19b): C30H28N2O5; MW = 496.56 g/mol;
0.388g (78.1%); white solid; mp 232-234 °C. H NMR (400
MHz, DMSO-d6) δ : 1.69-1.86 (2m, 4H, H-3/6, eq:ax),
2.15–2.28 (m, 4H, H-2/7), 2.43-2.58 (m, 4H, H-4/5, eq:ax),
3.71 (s, 3H, 3’-OCH3), 4.89 (s, 1H, H-9), 5.19 (s, 2H, H-1”),
1
Acknowledgments
6.61 (dd,
6.89 (d,
J
= 8.3, 1.9, 1H, H-6’), 6.85 (d,
J = 1.9, 1H, H-2’),
J
= 8.3, 1H, H-5’), 7.12 (s, 1H, H-4”’), 7.52 (m, 3H,
Thanks are due to FCT/MEC for the financial support for
the project PT-DZ/0005, of the QOPNA research Unit (FCT
UID/QUI/00062/2019) and CICECO – Aveiro Institute of
Materials (FCT Ref. UID/CTM/50011/2019) through
national founds and, where applicable, co-financed by the
FEDER, within the PT2020 Partnership Agreement, and to
the Portuguese NMR Network. We would like to thank the
General Directorate for Scientific Research and
Technological Development – DGRSDT of Algeria and
Agence Thématique de Recherche en Sciences et
Technologie ATRST for approving the co-financed bilateral
project PT-DZ/0005. We further wish to thank CICECO for
funding the purchase of the single-crystal X-ray
diffractometer. The authors are thankful to Pr. Farid
Messelmi the Dean of Faculty of Exact Sciences and
informatics in Djelfa University, for providing necessary
H-4””,H-3””/5””), 7.88 (m, 2H, H-2””/6””), 9.39 (s, 1H, 10-
NH) ppm. 13C NMR (100 MHz, DMSO-d6) δ(ppm): 21.4 (C-
3/6), 26.8 (C-4/5), 31.7 (C-9), 37.2 (C-2/7), 55.9 (3’-OCH3),
62.0 (C-1”), 102.7 (C-4”’), 112.8 (C-8a/9a), 112.9 (C-2’),
114.7 (C-5’), 119.4 (C-6’), 127.1 (C-2””/6””), 128.8 (C-
1””), 129.6 (C-3””/5””), 130.8 (C-4””), 142.1 (C-1’), 145.4
(C-4’), 149.0 (C-3’), 151.7 (C-4a/C-10a), 162.4 (C-3”’),
169.4 (C-5”’), 195.4 (C-1/8, C=O) ppm. HRMS (ESI+): m/z
calcd for [C30H28N2O5+H]+: 497.1998; found: 497.2071.
9-{4-[(3-phenylisoxazol-5-yl)methoxy]phenyl}-3,4,5,6,7,9-
hexahydro-1H-xanthene-1,8(2H)-dione (20a): C29H25NO5;
MW = 467.52 g/mol; 0.106g (75.5%); white solid; mp 238-
1
240 °C H NMR (400 MHz, DMSO-d6) δ: 1.74–2.01 (2m,
4H, H-3/6, eq:ax), 2.19–2.35 (m, 4H, H-2/7), 2.57 – 2.73 (m,
4H, H-4/5, eq:ax), 4.51–4.57 (s, 1H, H-9), 5.14–5.37 (s, 2H,
H-1”), 6.85–6.97 (d,
laboratory facilities
.
J
= 8.2 Hz, 2H, H-3’), 7.06–7.15 (d,
J
=
8.4 Hz, 2H, H-2’), 7.15–7.24 (s, 1H, H-4’”), 7.41–7.63 (q,
J
References and notes
= 3.2, 2.3 Hz, 3H, H-3””/4””), 7.78–7.96 (m, 2H, H-2””)
13
ppm. C NMR (100 MHz, DMSO) δ 20.2 (C-3/6), 26.9 (C-
1. (a) V. Dave, K. Tak, A. Sohgaura, A. Gupta, V. Sadhu, K. R.
Reddy. J. Microbiol. Methods 2019, 160, 130-142. (b) V. Dave,
A. Gupta, P. Singh, C. Gupta, V. Sadhu, K. R. Reddy. Nano-
Structures and Nano-Objects 2019, 18, 100288. (c) S. Gulla, D.
Lomada, V. V. S. S. Srikanth, M. V. Shankar, K. R. Reddy,
Sarvesh Soni, M. C. Reddy. Methods in Microbiology 2019, 46,
255-293. (d) C. Praveen Kumar, T. S. Reddy, P. S. Mainkar, V.
Bansal, R. Shukla, S. Chandrasekhar and H. M. Hügel, Eur. J.
Med. Chem., 2016, 108, 674–686.
4/5), 30.4 (C-9), 36.8 (C-2/7), 60.9 (C-1”), 102.6 (C-4”’),
114.6 (C-3’), 116.1 (C-8a/9a), 127.1 (C-2””), 128.8 (C-1””),
129.5 (C-3””), 129.6 (C-2’), 130.8 (C-4””), 138.2 (C-1’),
156.4 (C-4’), 162.4 (C-3”’), 165.1 (C-4a/10a ), 169.2 (C-
5”’), 196.8 (C-1/8, C=O). HRMS (ESI+): m/z calcd for
[C29H25NO5 +H]+: 468.1811; found: 468.1688.
Biological screening
2. (a) P. Wark, Respiratory Medicine, 2004, 98, 915–923. (b) Y. Hu,
C. Li, X. Wang, Y. Yang, H. Zhu, Chem. Rev., 2014, 114, 5572–

10
5610. (c) S. A. Steiger, C. Li, D. S. Backos, P. Reigan, N.R.
Natale, Bioorg. Med. Chem., 2017, 12, 3223-3234.
3. (a) D. Dheer, V. Singh, R. Shankar, Bioorg. Chem., 2017, 71, 30–
54. (b) Z.Najafi, M. Mahdavi, M. Saeedi, E. K. Razkenari, N.
Edraki, M. Sharifzadeh, M. Khanavi, T. Akbarzadeh, Bioorg.
Chem., 2019, 83, 303–316. (c) P. De Andrade, S. P. Mantoani, P.
Sérgio, G. Nunes, C. R. Magadán, C. Pérez, D. J. Xavier, E.
Tiemi, S. Hojo, N. E. Campillo, A. Martínez and I. Carvalho,
Bioorg. Med. Chem., 2019, 27, 931-943.
30. L. Saidi, D. H. A. Rocha, O. Talhi, Y. Bentarzi, B.-Nedjar-
Kolli, K. Bachari, F. A. A. Paz, L. A. Helguero, A. M. S.
Silva, Chem. Med. Chem.
31. F. S. Behbahani, J. Tabeshpour, S. Mirzaei, S.
Golmakaniyoon, Z. Tayarani Najaran, A. Ghasemi, R.
Ghodsi, Arch. Pharm. Chem. Life Sci. 2019, 352:e1800307.
, 2019, 14, 1041-1048.
‐
32. S. Mahantia, S. Sunkaraa, R. Bhavanib, Synth. Comm. 2019
,
49, 1729-1740.
33. T. Lisboa, D. Silva, S. Duarte, R. Ferreira, C. Andrade, A. L.
Lopes, J. Ribeiro, D. Farias, R. Moura, M. Reis, K. Medeiros,
4. C. D. Hein, X.-M. Liu, D. Wang, Pharm. Res., 2008, 25, 2216–
2230.
H. Magalhães, M. Sobral, Molecules 2020, 25, 64.
5. S. K. Mamidyala and M. G. Finn, Chem. Soc. Rev., 2010, 39,
1252.
34. E. Catanzaroa, F. Seghettib, C. Calcabrinia, A. Rampab, S.
Gobbib, P. Sestilic, E. Turrinia, F. Maffeia, P. Hreliad, A.
6. S. Kantheti, R. Narayan, K. V. S. N. Raju, RSC Adv., 2015, 5,
3687–3708.
Bisib, F. Bellutib, C. Fimognari, Bioorg. Chem. 2019, 86,
538-549.
7. K. Chiotos, N. Vendetti, T. E. Zaoutis, J. Baddley, L. Ostrosky-
Zeichner, P. Pappas, B. T. Fisher, J. Antimicrob. Chemother.,
2016, 71, 3536–3539.
Supplementary Material
8. A. Singh, S. T. Saha, S. Perumal, M. Kaur, V. Kumar, ACS
Omega, 2018, 3, 1263–1268.
9. L. D. Bailey, R. V. Kalyana Sundaram, H. Li, C. Duffy, R. Aneja,
A. Rosemary Bastian, A. P. Holmes, K. Kamanna, A. A. Rashad,
Supplementary material is available [NMR data and spectra
plus crystallographic details].
I. Chaiken, ACS Chem. Biol., 2015, 10, 2861–2873.
10. K. Lal, P. Yadav, A. Kumar, A. Kumar, A. K. Paul, Bioorg.
Chem., 2018, 77, 236–244.
11. H. M. Faidallah, S. S. Panda, J. C. Serrano, A. S. Girgis, K. A.
Khan, K. A. Alamry, T. Therathanakorn, M. J. Meyers, F. M.
Sverdrup, C. S. Eickhoff, S. G. Getchell, A. R. Katritzky, Bioorg.
Med. Chem., 2016, 24, 3527–3539.
12. A. Anand, M. V. Kulkarni, S. D. Joshi, S. R. Dixit, Bioorg. Med.
Chem. Lett., 2016, 26, 4709–4713.
13. F. Harris, L. Pierpoint, Med Res Rev, 2012, 29, 1292–1327.
14. D. A. Talan, W. R. Mower, A. Krishnadasan, F. M. Abrahamian,
F. Lovecchio, D. J. Karras, M. T. Steele, R. E. Rothman, R.
Hoagland, G. J. Moran, N. Engl. J. Med., 2016, 374, 823–832.
15. (a) V. D. da Silva, B. M. de Faria, E. Colombo, L. Ascari, G. P. A.
Freitas, L. S. Flores, Y. Cordeiro, L. Romão, C. D. Buarque,
Bioorg. Chem., 2019, 83, 87–97. (b) K. S. Raju, S. AnkiReddy, G.
Sabitha, V. S. Krishna, D. Sriram, K. B. Reddy, S. R. Sagurthi,
Bioorg. Med. Chem. Lett., 2019, 29, 284–290. (c) S. Burra, V.
Voora, Ch. P. Rao, P. V. Kumar, R. K. Kancha, G. L. D.
Krupadanam, Bioorg. Med. Chem. Lett., 2017, 27, 4314–4318. (d)
N. G. Khaligh, T. Mihankhah, M. R. Johan, J. Mol. Liq., 2019,
277, 794-804.
16. (a) F. Bossert, W. Vater, Med. Res. Rev., 1989, 9, 291–324. (b) S.
M. Derayea, D. M. Nagy, Rev. Anal. Chem., 2018, 37, 1-14. (c)
A. Kumar, K. M. Rao, S. S. Han, Carbohydr. Polym., 2018, 180,
128-144. (d) L. Hellen, S. Matos, F. T. Masson, L. A. Simeoni, M.
Homem-de-mello, Eur. J. Med. Chem., 2018, 143, 1779-1789.
17. D. J. Triggle, Cell Mol. Neurobiol., 2003, 23, 293–303.
18. S. Sivapalarajah, M. Krishnakumar, H. Bickerstaffe, Y. Y. Chan,
J. Clarkson, A. Hampden-Martin, A. Mirza, M. Tanti, A. Marson,
M. Pirmohamed, N. Mirza, Epilepsia, 2018, 59, 492–501.
19. A. G. Banerjee, L. P. Kothapalli, P. A. Sharma, A. B. Thomas, R.
K. Nanda, S. K. Shrivastava and V. V. Khatanglekar, Arab. J.
Chem., 2016, 9, S480–S489.
20. K. R. M. Naidu, B. S. Krishna, M. A. Kumar, P. Arulselvan, S. I.
Khalivulla, O. Lasekan, Molecules, 2012, 17, 7543–7555.
21. S. Naseem, M. Khalid, M. N. Tahir, M. A. Halim, A. A. C. Braga,
M. M. Naseer, Z. Shafiq, J. Mol. Struct., 2017, 1143, 235–244.
22. J. R. Kim, S. Michielsen, J. Photochem. Photobiol., 2015, 150,
50–59.
23. G. Harichandran, S. D. Amalraj, P. Shanmugam, J. Mol. Catal. A
Chem., 2014, 392, 31–38.
24. P. Tiposoth, S. Khamsakhon, N. Ketsub, T. Pongtharangkul, I.
Takashima, A. Ojida, I. Hamachi, J. Wongkongkatep, Sensor
Actuat B-Chem., 2015, 209, 606–612.
25. L. Malfatti, K. Suzuki, A. Erker, Y. Jang, P. Innocenzi, J.
Photochem. Photobiol., 2018, 357, 30–35.
26. A. Djemoui, M. R. Ouahrani, A. Naouri, L. Souli, S. Rahmani, M.
B. Lahrech, Heterocycl. Lett., 2018, 8, 455-467.
27. X. Creary, A. Anderson, C. Brophy, F. Crowell, Z. Funk, J. Org.
Chem., 2012, 77, 8756–8761.
28. A. Naouri, A. Djemoui, M. B. Lahrech, S. Rahmani and M. R.
Ouahrani, Heterocycl. Lett., 2017, 7, 589-597.
29. J. Grell, J. Bernstein, G. Tinhofer, Acta Cryst. B, 1999, 55, 1030-
1043.

Highlights:
•
1,2,3-triazole-acridinedione/xanthenedione
and
1,2-isoxazole-
acridinedione/xanthenedione heterocyclic hybrids have been synthesized via
1,3-dipolar coupling reaction
•
•
1D, 2D NMR and single-crystal X-ray diffraction analysis confirmed the hybrid
structure.
Anti-proliferative potential in breast and prostate cancer cell lines have been
evidenced for the O-1,2,3-triazole-xanthenedione hybrid molecules.

Declaration of interests
The authors declare that they have no known competing financial interests or personal relationships
that could have appeared to influence the work reported in this paper.