Novel Linezolid analogues with antiparasitic activity against Hymenolepis nana

Article abstract of DOI:10.1016/j.bioorg.2020.104359

The stereoselective synthesis and anti- Hymenolepis nana activity of six Linezolid-type compounds, obtained by chemical modification of L-Alanine, are reported in this work. The synthetic strategy was to prepare diasteromeric N,N-dibenzylamino oxazolidino

Full text of DOI:10.1016/j.bioorg.2020.104359

Journal Pre-proofs
Novel Linezolid Analogues with Antiparasitic Activity against Hymenolepis
nana
Eleazar Alcántar-Zavala, Esteban Hernández-Guevara, Adrián Ochoa-Terán,
Julio Montes-Ávila, Edgar A. Estrada-Zavala, Alex J. Salazar-Medina, Efraín
Alday, Alberto Cabrera, Gerardo Aguirre, Valentín Miranda-Soto, Carlos
Velazquez, Sylvia P. Díaz-Camacho, José L. Medina-Franco
PII:
S0045-2068(20)31657-6
DOI:
https://doi.org/10.1016/j.bioorg.2020.104359
YBIOO 104359
Reference:
To appear in:
Bioorganic Chemistry
Received Date:
Revised Date:
Accepted Date:
3 September 2020
22 September 2020
5 October 2020
Please cite this article as: E. Alcántar-Zavala, E. Hernández-Guevara, A. Ochoa-Terán, J. Montes-Ávila, E.A.
Estrada-Zavala, A.J. Salazar-Medina, E. Alday, A. Cabrera, G. Aguirre, V. Miranda-Soto, C. Velazquez, S.P.
Díaz-Camacho, J.L. Medina-Franco, Novel Linezolid Analogues with Antiparasitic Activity against Hymenolepis
nana, Bioorganic Chemistry (2020), doi: https://doi.org/10.1016/j.bioorg.2020.104359
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 Elsevier Inc. All rights reserved.

Novel Linezolid Analogues with Antiparasitic Activity
against Hymenolepis nana
1
2
1
Eleazar Alcántar-Zavala, Esteban Hernández-Guevara, Adrián Ochoa-Terán, * Julio
3
3
4
5
Montes-Ávila, * Edgar A. Estrada-Zavala, Alex J. Salazar-Medina, Efraín Alday,
1
1
1
5
Alberto Cabrera, Gerardo Aguirre, Valentín Miranda-Soto, Carlos Velazquez, Sylvia P.
6
7
Díaz-Camacho, José L. Medina-Franco
1
Centro de Graduados e Investigación en Química, Tecnológico Nacional de México/IT
de Tijuana. Tijuana, B.C., México.
2
Facultad de Ciencias Químicas e Ingeniería, Universidad Autónoma de Baja California
campus Tijuana. Tijuana, B. C. México.
3
Facultad de Ciencias Químico Biológicas, Universidad Autónoma de Sinaloa. Culiacán,
Sin. México.
4
Cátedras Conacyt - Departamento de Investigación en Polímeros y Materiales,
Universidad de Sonora. Hermosillo, Sonora, México.
5
Departamento de Ciencias Químico-Biológicas, Universidad de Sonora. Hermosillo,
Sonora, México.
6
Environment and Health Research Unit, Autonomous University of Occident, Los
Mochis, Sinaloa, México.
7
Departamento de Farmacia, Facultad de Química, Universidad Nacional Autónoma de
México. Ciudad de México, México.
*
Correspondence author e-mail: ochoa@tectijuana.mx, jmontes@uas.edu.mx
Abstract
The stereoselective synthesis and anti- Hymenolepis nana activity of six Linezolid-type
compounds, obtained by chemical modification of L-Alanine, are reported in this work.
The synthetic strategy was to prepare diasteromeric N,N-dibenzylamino oxazolidinones 1
and 2, and coupling with 4-(4-bromophenyl)morpholine (3) to obtain N,N-dibenzylamino
1

Linezolid analogues 4 and 5. A hydrogenolysis reaction over 4 and 5 resulted in amino-
free Linezolid analogues 6 and 7, which were acetylated to reach diasteromeric Linezolid
analogues 8 and 9. The six Linezolid analogues 4-9 show in vitro antiparasitic activity
against Hymenolepis nana cestode, but not against several bacterial strains. Interestingly,
compounds 6, 7 and 9 exhibit high potency, having shorter paralysis and death times after
exposure (6 to 10 and 18 to 21 min, respectively), shorter than those found with
antihelmintic compound Praziquantel (20 and 30 min) at 20 mg/mL. In addition, a
cytocompatibility assay of 6-9 with human cells (ARPE-19 cells) demonstrate a non-
cytotoxic effect at 0.4 mM. These results show the pharmacological potential of the newly
reported Linezolid-type analogues as antiparasitic agents against Hymenolepis nana.
Keywords: Linezolid analogues; antiparasitic activity; stereoselective synthesis
1
. Introduction
Intestinal parasitosis is a public health problem which affect 60 % of worldwide
1
population. Among a wide number of well-known parasite organisms, Hymenolepis nana
(H. nana) and Hymenolepis diminuta cestodes are recognized as the principal
hymenolepiasis infective agents. Hymenolepiasis is consider as a neglected disease
2
affecting about 175 million of people around the world. H. nana is a parasite capable to
complete its biological cycle inside a single host. Although this infection is not fatal, it
3
does have significant morbidity. This is a cosmopolite disease, but a higher prevalence is
present in countries with a tropical climate, affecting mainly school age children, elderly
and immunocompromised adults. In addition, there is a higher prevalence in the lower
income population countries, and it is associated to bad hygiene habits. Commonly, this is
an asymptomatic illness, but its clinical manifestations are diarrhea, stomachache,
3
-6
headache, weight loss and poor nutrient absorption. Praziquantel is the drug traditionally
used to treat intestinal or tissue cestodial parasitosis. However, recent studies have
revealed resistance of H. nana to praziquantel, as well as hepatotoxic, genotoxic and
6
2+
carcinogenic side effects. Calcium ion (Ca ) channels are the only moiety identified as
the molecular target of Praziquantel by indirect evidence, showing high affinity for
2

membrane receptors, affecting the cellular membrane permeability, promoting the fast Ca2+
absorption and inducing damages in the tegument of some parasites.7,8
Linezolid is the first commercially available antibiotic with a 1,3-oxazolidin-2-one
structural base. It was first synthesized in 1990 by Pharmacia-Upjohn laboratories (now
Pfizer) and approved by the FDA in 2000. This drug is used for treatment of nosocomial
infections caused by multi-resistant Gram positive bacterial strains as methicillin-resistant
Staphylococcus aureus (MRSA), penicillin-resistant Staphylococcus epiderminis (PRSE),
penicillin-resistant
Spreptococcus
pneumoniae
(PRSP),
vancomycin-resistant
Enterococcus faecium and Enterococcus faecalis (VREF).9-13 In addition, the activity of
1
4
Linezolid against Mycobacterium tuberculosis has been proved. This drug inhibits the
initiation and translation phases in the synthesis of bacterial proteins by intercalation
between the 30S and 50S ribosomal subunits.15,16 Linezolid structure consists in of
oxazolidinone ring (A ring), a fluorophenyl ring (B ring), a morpholine ring (C ring) and
an acetamide methyl substituent with a specific S configuration in C5 position of the A ring
(
Figure 1).17
Oxazole compounds have been described as bioactive molecules valuable for medicinal
chemistry, showing a broad spectrum of biological activities, including antiparasitic
1
8
properties. However, there are few studies in the literature focused on the antiparasitic
activity of Linezolid or analogues. These reports conclude in null activity, which justifies
its exclusive use as an antibacterial agent.19,20
Previously, we reported the stereoselective synthesis and antibacterial activity of a series
of N,N-dibenzylamino 1,3-oxazolidin-2-ones prepared by chemical modification of -
amino acids. The oxazolidinone derived from L-alanine 1 showed significant activity
against Methicillin Resistant Staphylococcus aureus (MRSA) isolated strains.21,22 In this
work, by using diasteromeric oxazolidinones 1 and 2 as precursors, we report the synthesis
of six new analogues having the characteristic structure of Linezolid (A, B and C rings)
with some modifications such as a) the presence of a chiral center and a methyl group
attached to the ring A substituent, b) variation in the configuration of the oxazolidinone
chiral center (S or R) and c) different substituents in the amino group (benzyl, hydrogen or
acetyl) (Figure 1). Remarkable results were found in antiparasitic activity assays,
suggesting the anti- H. nana potential of these oxazolidinone compounds. To the best of
3

our knowledge, this is the first report of Linezolid-related compounds with antiparasitic
activity.
O
O
H
N
N
O
N
O
F
Linezolid
O
N
O
N
O
O
O
O
N
O
N
O
N
N
O
O
NH
F
NBn2
F
NH2
F
4
6
8
O
N
O
N
O
O
O
O
N
O
N
O
N
N
O
O
NH
F
NBn2
F
NH2
F
5
7
9
Figure 1. Linezolid and Linezolid-analogues structure.
2
. Experimental
2
.1 Chemical synthesis
N-aryl coupling of oxazolidinones 1 and 2 with 4-(4-bromo-2- fluorophenyl)morpholine
3).
To a solution of the oxazolidinone (1 or 2)21,22 (0.32 moles), CuI (0.01 mmol), K CO (0.53
(
2
3
mmol), 4-(4-bromo-2-fluorphenyl)morpholine (3) in toluene (25 mL) was added dropwise
the N,N´-dimethylmethanediamine (0.06 mmol). Then, the reaction mixture was stirred at
1
15°C for 72 h. Finally, water (25 mL) was added to finish the reaction and the organic
phase was separated, filtered using zeolite and evaporated under vacuum. The product
purification was carried out by flash chromatography on silica gel and a mobile phase of
petroleum ether and ethyl acetate (1:1).23,24
(R)-5-((S)-1-(dibenzylamino)ethyl)-3-(3-fluoro-4-morpholinophenyl)oxazolidin-2-one
(4). White solid, yield 75%. M. P. 68.5-69.5 °C FTIR: 3059, 2955, 1753, 1514, 1449,
-1 1
1
402, 1234, 1117 cm . H NMR (400 MHz, CDCl ): δ 7.30 (m, 10 H), 7.18 (dd, J = 11.8,
3
4

2
.5 Hz, 1H), 7.00 (ddd, J = 7.1, 2.5, 1.0 Hz, 1H), 6.89 (dd, J = 9.1, 9.0 Hz, 1H), 4.40 (ddd,
J = 8.3, 8.2, 7.2 Hz, 1H), 3.86 (m, 4H), 3.80 (t, J = 8.7 Hz, 1H), 3.75(d, J = 13.5 Hz, 2H),
.57 (dd, J = 9.0, 6.6 Hz, 1H), 3.48 (d, J = 13.5 Hz, 2H), 3.08 (m, 4H) 2.88 (m, 1H), 1.30(d,
3
1
3
J = 6.6 Hz, 3H). C NMR (100 MHz, CDCl ): δ 155.5 (d, J = 244.4 Hz), 154.6, 139.1,
3
C-F
1
36.2 (d, JC-F = 9.0 Hz), 133.3 (d, JC-F = 10.5 Hz), 128.9, 128.5, 127.4, 118.7 (d, JC-F = 4.1
Hz), 114.2 (d, JC-F = 3.3 Hz), 107.7 (d, JC-F = 25.8 Hz), 74.2, 66.9, 56.7, 54.6, 51.9, 49.4,
+
8
.6. HRMS (FAB ) calculated for C H FN O: 490.2506. Found: 490.2513 uma.
2
9
33
3
((S)-5-((S)-1-(dibenzylamino)ethyl)-3-(3-fluoro-4-morpholinophenyl)oxazolidin-2-
one (5). White solid, yield 70%. M. P. 78.0-79.0 °C. FTIR: 3032, 2955, 1753, 1514, 1449,
-
1 1
1
7
402, 1234, 1117 cm . H NMR (400 MHz, CDCl ): δ 7.37 (dd, J = 11.8, 2.6 Hz, 1H),
3
.30 (m, 10 H), 7.00 (ddd, J = 7.3, 2.0, 1.3 Hz, 1H), 6.94 (dd, J = 9.2, 9.0 Hz, 1H), 4.56
(ddd, J = 5.4, 5.1, 5.1 Hz, 1H), 3.93 (d, J = 13.6 Hz, 2H), 3.88 (m, 4H), 3.68 (dd, J = 8.4,
2
.4 Hz, 1H), 3.48 (d, J = 13.6 Hz, 2H), 3.07 (m, 4H), 2.92 (dc, J = 6.8, 5.1 Hz, 1H), 1.26
1
3
(
d, J = 6.8 Hz, 3H). C NMR (100 MHz CDCl ): δ 155.5 (d, J = 244.5 Hz), 154.5, 139.6,
3 C-F
1
36.2 (d, JC-F = 8.9 Hz), 133.5 (d, JC-F = 10.5 Hz), 128.9, 128.3, 127.0, 118.7 (d, JC-F = 4.2
Hz), 113.7 (d, JC-F = 3.3 Hz), 107.3 (d, JC-F = 26.3), 76.2, 67.0, 55.1, 54.8, 51.0, 47.6, 10.2.
+
HRMS (FAB ) calculated for C H FN O: 490.2506. Found: 490.2513.
2
9
33
3
Hydrogenolysis of 4 and 5
Compounds 4 or 5 (1 g, 2.04 mmol) were dissolved in dry methanol (25 mL) and Pd/C
0% (0.02 g, 0.20 mmol) was added. Then two drops of acetic acid were added, and the
1
reaction mixture was stirred for 4 h under H atmosphere. Then, the mixture was filtered
2
on a Celite pad in order to remove the Pd/C, and the organic solvent was evaporated under
vacuum conditions.
(R)-5-((S)-1-aminoethyl)-3-(3-fluoro-4-morpholinphenyl)oxazolidin-2-one (6). White
solid, yield 90%. M. P. 196-197 C. FTIR: 3373, 2952, 2835, 1740, 1636, 1514, 1445,
-
1 1
1
7
404, 1237, 1112 cm . H NMR (400 MHz, CDCl ): δ 7.50 (dd, J = 11.8, 2.6 Hz, 1H),
3
.22 (ddd, J = 8.8, 1.5, 1.2 Hz, 1H), 7.08 (dd, J = 9.1, 9.0 Hz, 1H), 4.77 (m, 1H), 4.10 (t, J
5

=
9.3 Hz, 1H), 4.00 (m, 1H), 3.74 (m, 4H), 3.40 (m, 1H), 2.96 (m, 4H) 1.16 (d, J = 6.6 Hz,
1
3
3
H). C NMR (100 MHz, CDCl ): δ 155.5 (d, J = 220.4 Hz), 154.3, 136.2 (d, JC-F = 7.3
3 C-F
Hz), 133.6 (d, JC-F = 10.2 Hz), 119.7 (d, JC-F = 4.1 Hz), 114.8 (d, JC-F = 3.3 Hz), 107.3 (d,
+
2
3
6.1), 74.2, 66.6, 51.1, 48.7, 46.5, 14.6. HRMS (FAB ) calculated for C H FN O :
1
5
21
3
3
10.1567. Found: 310.1621.
(S)-5-((S)-1-aminoethyl)-3-(3-fluoro-4-morpholinophenyl)oxazolidin-2-one
(7).
White solid, 80% yield. M. P. 128-129 C. FTIR: 3373, 2956, 1744, 1625, 1514, 1407,
-1 1
1
234, 1114 cm . H NMR (400 MHz, CDCl ): δ 7.51 (dd, J = 12.4, 2.6 Hz, 1H), 7.23 (dd,
3
J = 8.8, 1.9 Hz, 1H), 7.05 (dd, J = 9.8, 9.1Hz, 1H), 4.45 (ddd, J = 8.7, 8.6, 7.8 Hz, 1H),
4
2
2
.01 (dd, J = 9.0, 8.9 Hz, 1H), 3.83 (dd, J = 9.0, 8.9 Hz, 1H), 3.73 (m, 4H), 3.25 (bs, 2H),
1
3
.96 (m, 5H), 1.03 (d, J = 6.5 Hz, 3H). C NMR (100 MHz, CDCl ): δ 155.5 (d, J
=
3
C-F
42.1 Hz), 154.7, 135.8 (d, JC-F = 8.8 Hz), 134.1 (d, JC-F = 10.4 Hz), 119.7 (d, JC-F = 4.2
Hz), 114.4 (d, JC-F = 3.2 Hz), 107.3 (d, JC-F = 26.0Hz), 75.8, 66.6, 51.2, 49.2, 47.1, 18.2.
+
HRMS (FAB ) calculated for C H FN O : 310.1567. Found: 310.1621.
1
5
21
3
3
N-acetylation of 6 and 7
To a solution of 6 or 7 (1 g, 3.23 mmol) in dry dichloromethane (50 mL), TEA (0.1 mL,
.5 mmol) was added dropwise at -78 C under argon atmosphere and the mixture was
6
stirred for 15 min. Then a solution of acetyl chloride (0.38 g, 4.85 mmol) in 5 mL of solvent
was added dropwise, and the reaction was stirred for 45 min at the same temperature. Then
the organic layer was washed with brine, separated, dried over Na SO and filtered. The
2
4
solvent was evaporated under reduced pressure.
N-((S)-1-((R)-3-(3-fluoro-4-morpholinphenyl)-2-oxooxazolidin-5-il)ethyl)acetamide
(8). Yellow solid, 76% yield. M. P. 140-142 C. FTIR: 3323, 3063, 2920, 1741, 1659,
-1 1
1
514, 1220 cm . H NMR (400 MHz, CDCl ): δ 7.45 (dd, J = 15.0, 2.5 Hz, 1H), 7.13 (dd,
3
J = 9.1, 2.5 Hz, 1H), 6.93 (t, J = 9.2 Hz, 1H), 6.02 (d, J = 8.4 Hz, 1H), 4.70 (m, 1H), 4.25
(m, 1H), 4.08 (t, J = 9.1 Hz, 4H), 3.88 (m, 4H), 3.78 (dd, J = 9.0, 8.8 Hz, 1H), 3.04 (m,
1
3
4
H), 2.00 (s, 1H), 1.25 (d, J = 7.0 Hz, 3H). C NMR (100 MHz, CDCl ): δ 169.9, 155.0
3
6

(d, JC-F = 242.2 Hz), 154.4, 136.5 (d, JC-F = 8.8 Hz), 132.8 (d, JC-F = 10.5 Hz), 118.7 (d, JC-F
=
4.1 Hz), 114.0 (d, JC-F = 3.0 Hz), 107.5 (d, JC-F = 26.0Hz), 74.4, 66.9, 51.0, 47.7, 47.6,
+
2
3.2, 13.4. HRMS (FAB ) calculated for C H FN O : 352.1673. Found: 352.1705.
1
7
23
3
4
N-((S)-1-((S)-3-(3-fluoro-4-morpholinphenyl)-2-oxooxazolidin-5-il)ethyl)acetamide
(9). White solid, 70% yield. M. P. 196-198 C. FTIR: 3323, 3063, 2920, 1741, 1659, 1514,
-
1 1
1
8
4
=
220 cm . H NMR (400 MHz, CDCl ): δ 7.42 (dd, J = 15.0, 2.5 Hz, 1H), 7.05 (dd, J =
3
.8, 2.5 Hz, 1H), 6.90 (dd, J = 9.26, 9.0 Hz, 1H), 6.41 (d, J = 8.0 Hz, 1H), 4.60 (m, 1H),
.45 (m, 1H), 3.94 (t, J = 9.0 Hz, 4H), 3.88 (m, 5H), 3.05 (m, 4H), 1.97 (s, 1H), 1.35 (d, J
1
3
7.0 Hz, 3H). C NMR (100 MHz, CDCl ): δ 170.9, 155.4 (d, J = 245.0 Hz), 154.6,
3
C-F
1
3
36.4 (d, JC-F = 8.7 Hz), 132.8 (d, JC-F = 10.5 Hz), 118.7 (d, JC-F = 4.2 Hz), 113.9 (d, JC-F
=
.0 Hz), 107.5 (d, JC-F = 26.0Hz), 75.6, 66.9, 51.0, 47.6, 46.2, 22.1, 17.8. HRMS (FAB+)
calculated for C H FN O : 352.1673. Found: 352.1705.
1
7
23
3
4
2
.2 Antibacterial activity of Linezolid analogues
The antibacterial activity of all molecules was evaluated against twelve bacterial strains
eleven Gram-positive and one Gram-negative, see Table S7).25-27 All bacteria were
(
cultured in Muller-Hinton broth. MIC of all compounds were determined by broth
microdilution method according to CLSI guidelines. The 18-20 h grown culture gives about
1
08 CFU/mL of bacteria. The bacterial cultures were diluted to give approximately 106
CFU/mL in Muller-Hinton broth media and then were used to determinate antibacterial
activity.
The evaluation of the antibacterial activity of all compounds against different Gram-
positive and Gram-negative strains was performed at 100 μg/mL into 96-wells ELISA type
dishes. Then, the active compounds were evaluated at lower concentration in the range of
5
0 to 1.17 g/mL.
2
.3 In vitro antiparasitic activity of Linezolid analogues
Antiparasitary evaluations were made at one concentration that corresponds to the
Praziquantel solubility (20 mg/mL) in Hank’s medium. Hymenolepis nana worms were
7

exposed to Linezolid analogues or Praziquantel (positive control) in 24-well sterile plates,
worms/well in 1 mL of medium. Parasites in Hank’s medium were used as negative
control. The plates were monitored by light microscope every hour up to 12 h and then
5
2
8
every 24 h up to 72 h; morphology, mobility and vitality were registered. Concentration-
response assays of selected Linezolid analogues (10, 15 and 20 mg/mL) on the antiparasitic
activity were performed.
2
.4 Cellular viability assay
Insight about the effect of the linezolid analogues 6 to 9 on cell proliferation, cell viability
was assessed on human retinal pigment epithelial cell line (ARPE-19) by MTT assays29
3
0
with some modifications. ARPE-19 (CRL-2302) were purchased from American Type
4
Culture Collection (ATCC). Initially, ARPE cells (1×10 per well, 50 µL) maintained in
®
DMEM (Sigma-Aldrich) supplemented with 5% FBS (Gibco , Thermo Fisher Scientific),
®
were placed in each well of a 96-well plate (Costar , USA). The analogues were dissolved
in dimethyl sulfoxide (DMSO) at 40 mM. After 24 h of incubation at 37 °C in a 5% CO2
atmosphere to allow cells attachment, aliquots (50 µL) of the medium containing different
concentrations (12.5 to 400 µM) of linezolid analogues were added to cell cultures, and
were incubated for 48 h. We used the cytotoxic drug Doxorubicin hydrochloride (Sigma-
Aldrich) as a positive control in cellular viability assays. In the last 4 h of each assay, 10
µL of an MTT (Thiazolyl Blue Tetrazolium Bromide; Sigma-Aldrich) solution (5 mg/mL)
were added to each well. The cell viability was calculated by the ability of metabolically
active cells to reduce tetrazolium salt formazan. The formazan crystals were dissolved with
acidic isopropyl alcohol (0.3 %). The OD was measured on a microplate reader (Multiskan
EX, ThermoLabSystem) using a test wavelength of 570 nm and a reference wavelength
6
30 nm. The effect over cell viability of linezolid analogues was reported as a proliferation
percentage to each concentration evaluated.
8

3
. Results and discussion
3
.1 Chemistry
Oxazolidinones 1 and 2 were prepared following a methodology previously reported in 90
and 84% global yield.21,22 Compound 3 was obtained starting by a nucleophilic aromatic
substitution reaction of 3,4-difluoronitrobenzene and morpholine in a 90% yield. Then, the
nitro group was reduced under a H atmosphere using the Pd/C catalyst to obtain the amino
2
derivate in 97% yield. The amino group was exchange by a bromine with a 80% yield
under a Sandmeyer reaction conditions (Scheme S1).23,24 Aryl coupling of 3 with
oxazolidinones 1 and 2 leaded to N,N-dibenzylamine Linezolid analogues 4 and 5 in 75
and 70% yield, respectively. Then, a hydrogenolysis reaction allowed the synthesis of
amino-free derivates 6 and 7 in 90 and 80% yield. Finally, compounds 6 and 7 were
acetylated to form 8 and 9 in 70 and 76% yield, respectively (Scheme 1). Analogues 8 and
9
have high estructural resemblance to Linezolid, but with two significant differences, their
molecules have two chiral centers and a methyl group attached to the acetamide methyl
substituent.
Scheme 1. Synthetic route for the preparation of Linezolid analogues.
F
O
O
N
F
F
O
O
H
Br
N
O
NH
N
N
3
O
O
NBn2
CuI, K2CO3
MeNH(CH2)2NHMe
Toluene, 24 h
Bn2N
4
5
1
O
O
N
O
O
NH
NBn2
NBn2
2
H2,
Pd/C
MeOH
O
N
O
F
F
O
O
N
N
N
O
O
O
O
NHAc
NH2
CH3COCl
6
8
9
TEA
DCM
O
N
O
N
F
F
O
O
N
N
NHAc
NH2
7
9

1
13
1
All compounds were characterized by FTIR, H and C NMR, and mass spectrometry. H
NMR spectrum of oxazolidinone 1 shows the characteristic signals of benzyl groups at
7
.30 ppm for aromatic hydrogens, and at 3.75 and 3.47 ppm two doublets assigned to
methylene hydrogens. There is a broad singlet at 5.46 ppm belonging to carbamate
hydrogen and a multiplet at 4.50 ppm assigned to oxazolidinone methine. The signals for
oxazolidinone methylene hydrogens appear as two triplets located at 3.58 and 3.25 ppm.
The signal of alanine methine (asymmetric center) is present at 2.86 ppm and there is a
doublet signal located at 1.26 assigned to methyl hydrogens (Figure S1). The oxazolidinone
1
3
carbonyl signal appears at 159.6 ppm in the C NMR spectrum of 1. There are four
aromatic signals from 139.3 to 127.3 ppm and benzylic methylene signal is located at 54.6
ppm. The oxazolidinone methine and methylene signals are observed at 78.5 and 44.8 ppm,
respectively. The signal of alanine asymmetric carbon is located at 56.7 ppm and methyl
carbon at 8.7 ppm (Figure S2). A stretching vibration band for carbonyl is located at 1746
cm-1 in the FTIR spectrum (Figure S3). The MS(ESI) analysis of 1 shows the
+
+
+
quasimolecular ion [M+H] with 311 amu and adducts [M+Na] and [M+K] with 333 and
49 amu, respectively (Figure S4). The NMR spectra of 2 show the same signals than 1,
3
but slight differences in their chemical shifts are observed due to the different configuration
at C5 position (Figures S5 and S6). Oxazolidinone 1 crystallized as a dimer in the
monoclinic crystalline system with a space group P2 (Figure 2 and Table S1).
1
Interestingly, the asymmetric unit of 1 showed two oxazolidinone molecules connected to
each other by hydrogen bonding of oxazolidinone unit. These two intermolecular hydrogen
bonds (NHO=C) have been described as a strong non-covalent interaction with
DonorAcceptor distances of 2.889 and 2.939 Å.31
1
0

Figure 2. Dimeric molecular structure of compound 1 with thermal ellipsoids drawn at
0% probability level. Bond distances of the non-covalent interactions DA (Å):
2
N(3)O(1)=C, 2.939(4); N(1)O(3)=C, 2.889(4).
In addition, the hydrogens of chiral carbons (C1 and C4) have an anti-conformation with
an R and S absolute configuration on C1 and C4, respectively (Flack parameter = 0.05(8).
The crystal arrangement exhibits intermolecular C–HO hydrogen bonds, which together
with the intermolecular hydrogen bonds of the type NHO=C, led to formation of the 2D
framework (Figure S7).
1
The H NMR spectrum of 3 shows a multiplet at 7.26 ppm and a triplet a 6.89 ppm
corresponding to aromatic hydrogens. There are two multiplets at 3.87 and 3.07 ppm
belonging to morpholine fragment (Figure S8). Some aromatic signals located at 156.7 to
1
3
1
13.9 ppm in the C NMR spectrum present a doublet multiplicity due to coupling with
fluorine. The characteristic signal for the ipso fluorine carbon is located at 155 ppm with a
J= 249.3 Hz. The aliphatic carbons are located at 66.9 and 50.8 ppm, respectively (Figure
S9). In addition, it was possible to obtain the X ray crystal structure of 3 (Figure 3, Table
S2) and its synthetic precursors 12 and 13 (Figure S10, Tables S3 and S4), which have not
been published. The molecular structure of 3 and its precursors have morpholine ring in a
chair-conformation with nitrogen substituted at the equatorial position by the aryl group
(Figure 3). The crystal arrangement exhibits intermolecular C–H···F and C–H···O
hydrogen bonds in compound 3. Moreover, the crystal of 3 presents one π-π interaction
between the phenyl rings of two molecules (Figure S11).
1
1

Figure 3. Molecular structure of compound 3 with thermal ellipsoids drawn at 50%
probability level.
1H NMR spectra of 4 and 5 show signals corresponding to both oxazolidinone and
fluorophenylmorpholine moieties, with significant differences in the chemical shift of
oxazolidinone methylene and fluorophenyl hydrogens. The rest of signals have similar
chemical shifts to their respective precursors (Figures S12 and S14). Similarly, all carbon
signals belonging to both fragments are presented, but significant differences in the
chemical shift of oxazolidinone carbons C4 and C5, morpholine C14 and fluorophenyl C8
signals are observed, due to the coupling of both fragments (Figures S13 and S15). The
-1
FTIR spectra of 4 and 5 show a stretching band for carbonyl at 1744 cm (Figure S16).
+
The MS(ESI) analysis of these compounds show a quasimolecular ion [M+H] with 490
+
amu and the adduct [M+Na] with 512 amu (Figure S17).
1
Signals belonging to benzylic groups are no longer present in the H NMR spectra of 6 and
7
, and oxazolidinone signals are shifted downfield (Figures S18 and S20). Only the
1
3
fluorphenyl signals are present in the aromatic zone in C NMR spectra, the rest of
aliphatic carbon signals have different chemical shift (Figures S19 and S21). FTIR spectra
-1
show the amino-group stretching vibrations at 3384 y 3365 cm , and carbonyl stretching
-1
+
at 1734 cm (Figure S22). The MS(ESI) analysis show a quasimolecular ion [M+H] with
10 amu corresponding to these compounds (Figure S23).
3
1H NMR spectra of acetylated derivates 8 and 9 present two new signals due to the acetyl
group. There is a doublet at 6 ppm and a singlet at 2 ppm assigned to amide and a methyl
1
2

hydrogens, respectively. The chemical environment of the rest of hydrogens was affected
by the presence of acetyl group, particularly the H6 signal shifted 0.8 and 1.4 ppm in 8 and
1
3
9
, respectively (Figures S24 and S26). C NMR spectra show the new carbonyl and methyl
signals at 170 and 23 ppm, respectively (Figures S25 and S27). There are two new
-1
stretching bands at 3275 and 1647 cm in FTIR spectra due to the presence of acetyl group
in these molecules (Figure S28). The MS(ESI) analysis of 8 and 9 shows a quasimolecular
+
+
+
ion [M+H] with 352 amu, and adducts [M+Na] and [M+K] with 374 and 391 amu,
1
respectively (Figure S29). Figure 4 shows the sequence of H NMR spectra obtained from
the oxazolidinone 1 to Linezolid analogue 8. A full assignation of NMR signals is
summarized in Tables S5 and S6.
O
1
2
O
5
7
NH
3
6
a)
4
NBn2
1
O
2
4
9
F
10
1O
5
14 15
O
8
1
N
3
7
6
N
b)
c)
11
3
NBn2
12
4
O
2
9
F
10
1O
14 15
8
N
3
7
6
5
N
4
11
O
13
NH2
12
6
O
2
9
F
1O
5
14 15
O
8
1
10
N
7
6
3
N
11
d)
4
3
O
NH16
12
17
8
18
1
Figure 4. Partial H NMR spectra of 1 (a), 4 (b), 6 (c) and 8 (d).
3
.2 In vitro activity of Linezolid analogues against bacterial strains
First, the antibacterial activity of dibenzylamino oxazolidinones 1 and 2, and Linezolid
analogues 4-9 was evaluated against Gram-positive and Gram-negative strains (Table S7)
at 100 g/mL. Linezolid was evaluated as positive control against the same strains. The
1
3

active compounds at this concentration were 1, 2, 5 and 6, only against Gram positive
strains. Dibenzylamino oxazolidinone 1 was active against group A Streptococcus 01 and
ATCC 25923 Staphylococcus aureus and dibenzylamino oxazolidinone 2 was active
against group A Streptococcus 01, Staphylococcus aureus 05, ATCC 25923
Staphylococcus aureus, MRSA-01, MRSA-03 and MRSA-05. Linezolid analogues 5 and
6
were active against Staphylococcus aureus 05.
Then, their antibacterial activity was evaluated at lower concentration in the range of 1.17
to 50.0 g/mL. The results showed that at lower concentration these compounds do not
display activity against all the initial strains. Dibenzylamino oxazolidinones 1 and 2 only
inhibited the growing of group A Streptococcus 01 strain, but a small bacterial button was
observed even at the highest concentration (50.0 g/mL) and the Linezolid analogues 5
and 6 showed activity only against Staphylococcus aureus 05 at 50 µg/mL.
The experimental results demonstrated the poor antibacterial activity of these Linezolid
analogues compared to Linezolid (Table S8). In order to rationalize this significant
difference in the antibacterial activity at the molecular level, the molecular docking of
Linezolid analogues 4-9 in the Phosphoryl Transferase Center (PTC) of Linezolid was done
3
2
using the MOE 2018 program. The docking was performed with the crystallographic
structure of ribosomal RNA (rRNA) of E. coli (Proten Data Bank ID: 4V4Q) with the same
protocol previously reported for another Linezolid analogues.33 The structure of all
3
4
Linezolid analogues, excepting 4, occupy the PTC of E. coli (Figures S30 to S36). The
protonated forms of 6 and 7 were also calculated. Table 1 shows the score and placement
energies (in kcal/mol) of Linezolid and analogues. The score value obtained after several
calculations is related to the quality of the coupling, where the lower scores indicate the
most favorable couplings. As can be seen, analogues 5, 6 and 8 have similar score than
Linezolid, but they do not show potent antibacterial activity as Linezolid. Some plausible
reasons for this difference could be: 1) a highly superior placement energy to Linezolid,
probably due to the steric effect of voluminous groups as benzyl or methyl present in their
molecular structure, and 2) different intermolecular interactions involved in the
supramolecular complexes. The docking of Linezolid in the PTC of E. coli shows the
1
4

typical hydrogen bond interaction between the amide hydrogen and the phosphodiester
group of G2505 nucleotide, which is the main interaction responsible of the antibacterial
activity.35 Linezolid analogue 5 presents a non-typical hydrogen bond with the
G2061acting as H-acceptor, while analogue 6 presents two interactions, one hydrogen bond
with G2505 acting as H-donor and a lower energy C-H∙∙∙ interaction with A2451 (Figure
5
). These analogues display poor activity against one bacterial strain. The rest of Linezolid
analogues which are totally inactive establish low energy interactions such as C-H∙∙∙ and
N-H∙∙∙, but no hydrogen bonds are present (Figures S30 to S36).
Table 1. Score and placement energy (kcal/mol) of Linezolid analogues.
Compound
Score
kcal/mol)
-7.20
Placement Energy
(kcal/mol)
(
Linezolid
5
-70.35
-7.18
-5.85
-7.18
-6.28
-6.69
-7.19
-6.88
-9.27
-41.51
-53.12
-46.24
-39.42
-54.64
-53.92
6
-H+
6
7
-H+
7
8
9
1
5

a)
b)
c)
d)
Figure 5. Molecular docking and interaction maps of 5 (a and b) and 6 (c and d) with the
Phosphoryl Transferase Center (PTC) of the crystallographic structure of ribosomal RNA
(rRNA) of E. coli.
3
.3 In vitro activity of Linezolid analogues against Hymenolepis nana
Based on results obtained in the antibacterial experiments, it was decided to evaluate the
in vitro antiparasitic activity of Linezolid analogues 4-9 against H. nana at 20 mg/mL
following a procedure previously reported by Montes-Ávila.28 The study consists of
observing the parasite mobility and morphology in the presence of Linezolid analogue, as
well as its viability using the Evans staining. Analogues 4 and 5 induced paralysis and
death at 20 and 60 min, respectively (Table 2). Death time was twice that observed with
1
6

Praziquantel (30 min), but interestingly, compounds 4 and 5 cause morphological damage
to the parasite’s scolex, neck and proglottids (Figure 6b). Analogues 6 and 7 have a similar
time of paralysis (11 and 10 min, respectively) and death time (21 min). Both compounds
showed higher potency than Praziquantel. The staining showed morphological damage to
the gravid proglottids, scolex and eggs caused by 6 (Figure 6c). These effects in the parasite
were not observed with Praziquantel and they are important due to the parasitosis
persistence is associated with the scolex attachment to the intestine and with the reinfection
by the eggs present in the gravid proglottids.36 Based on these results, the chemical
configuration in 4 and 5 does not have effect over the antiparasitic activity, as well as in 6
and 7. Paralysis and death times were similar with these diasteromeric pairs. Instead,
analogues 8 and 9 had significant differences in activity, the death time observed with 8
was three-times that of 9. Analogue 9 was the most active Linezolid analogue against H.
nana with a death time of almost half (18 min) found with Praziquantel (30 min). More
details of the anatomic damage caused by these compounds in the parasite are provided in
Table 2. Linezolid only caused a reversible paralysis in the worms, there was not anatomic
damage and the parasite was alive even after 24 h of treatment.
Table 2. Antiparasitic activity of Linezolid analogues against Hymenolepis nana at 20
mg/mL.
Paralysis Death
Compound
4
Structure
Anatomic effect
(min)
(min)
Partial staining, damage in
gravid proglottids,
moderate release of
unstained eggs
Partial staining, damage in
neck, scolex and mature
proglottids
O
F
O
N
N
20
60
O
NBn2
O
N
F
O
5
N
20
60
O
NBn2
Total staining,
O
N
F
F
O
damage in the gravid
proglottids, scolex and
release of stained eggs
6
7
N
N
11
10
21
21
O
O
NH2
O
N
O
Total staining
NH2
1
7

O
N
F
F
Partial staining, damage in
neck, scolex and mature
proglottids
O
NHAc
O
8
9
N
N
2
6
60
18
O
O
O
N
Total staining
Total staining
NHAc
Praziquantel
Linezolid
20
5
30
Reversible paralysis, no
anatomic damage
N/D
Normal mobility, no
anatomic damage
Control (-)
N/D
N/D
N/D indicates that parasites were still moving or alive up to 16 h.
a)
b)
c)
Figure 6. Pictures of H. nana worms a) without treatment, b) treated with 4 for 1 h and c)
treated with 6 for 20 min.
Antiparasitic activity of the most active analogues 6 to 9 was study at 20, 15 and 10 mg/mL
(Table 3). Paralysis and death times gradually increase at lower concentrations but, with
exception of 8, the antiparasitic activity of Linezolid analogues was preserved.
Interestingly, analogues 7 and 9 (S,S stereoisomers) were more active than 6 and 8 (R,S
stereoisomers). In fact, the activity of 8 diminished considerably at lower concentration.
The most active analogues at lower concentration were 6 and 7. Several factors may be
considered in the antiparasitic activity of these compounds, but the solubility in aqueous
media plays an important role. Amino-free (6 and 7) and acetylated (8 and 9) analogues
have higher solubility than 4 and 5, enhancing the availability to interact with the parasite.
1
8

Table 3. Antiparasitic activity of Linezolid analogues against Hymenolepis nana at
different concentration.
Time
Concentration
Compound
Structure
Paralysis
Death
(min)
120
40
21
60
(mg/mL)
(min)
O
F
10
15
20
10
20
15
11
15
O
N
6
7
8
9
N
O
O
NH2
O
N
F
O
N
N
15
10
60
NH2
2
0
10
10
10
2
21
N/D
N/D
60
O
N
10
15
F
F
O
O
NHAc
2
0
O
10
15
10
90
O
N
10
6
90
18
N
O
NHAc
2
0
N/D indicates that parasites were still moving or alive up to 16 h.
Considering the paralysis effect of Linezolid and its analogues over the parasite, it is
possible that their mechanism of action being the same as Praziquantel, that is, the
2
+
alteration of the Ca ions flux through the cell membrane caused by the interaction of these
compounds into the ions channels. In this regard, it is important to point out that although
Praziquantel and Linezolid do not have a similar structure, they share some similitudes
such as having an aromatic group, a heterocyclic ring and an exocyclic carbonyl group,
which presumably establish similar intermolecular interactions. On the other hand, the
potent activity and morphological damage showed by the Linezolid analogues may be
consequence of a stronger interaction due to the additional chiral center, as well as the
bulky groups present in their structures inducing conformational and tridimensional
arrangements favoring their interaction into the ions channels.
1
9

3
.4 Cellular viability assay
In order to evaluate whether the Linezolid analogues (6-9) induce changes over
proliferation profile of ARPE-19 cells, their effect on cell viability was evaluated by MTT
assay after 48 h of treatment.29,30 None of the analogues significantly affected the cell
viability at any concentration tested (50, 100, 200 and 400 µM) (Figure S37). After 48h
treatment, Linezolid analogues decreased cell proliferation between 5-20%,
approximately, at the highest concentration evaluated (400 µM), in comparison with
DMSO dissolvent control (Table 4). Unlike, Doxorubicin induced a strong antiproliferative
effect over ARPE-19 cells and reduces the cell proliferation in a 30% at 0.2 µM.
Table 4. Proliferation percentages of ARPE-19 cells culture with Linezolid analogues at
the highest concentration tested (400 µM)
Compound
6
Structure
Proliferation %a
O
F
O
N
N
76.5 ± 1.7
O
NH2
O
N
F
O
7
N
85.2 ± 1.0
94.9 ± 1.2
89.9 ± 2.3
83.1 ± 4.3c
O
NH2
O
N
F
F
O
8
9
N
N
O
O
NHAc
O
O
N
NHAc
O
N
F
O
H
N
Linezolid
N
O
O
a
b
c
Data is expressed as mean of three independent experiments performed by triplicate.
Doxorubicin was employed as a control.
Evaluated at 100 M.
ARPE-19 cells not only remained metabolically active after the treatment, but also
maintained their cellular integrity, indicating non-cytotoxic effects of Linezolid analogues
(Figure S38).
2
0

Finally, important molecular properties such as LogP , H-donor and H-acceptor number,
ow
molecular weight (MW), knowing as Lipinkiꞌs rule or the rule of 5,37,38 and total polar
surface area (TPSA) were calculated. The results predict these Linezolid analogues might
have good permeability after orall administration (Table S14).
4
. Conclusions
Herein, we present the first Linezolid type compounds reported with in vitro antiparasitic
activity against Hymenolepis nana. The biological evaluations demonstrate their promising
potential as antiparasitic agents because they are more active and induce anatomic damages
not observed with Praziquantel, a common drug used in the treatment of this parasitosis.
Noteworthy, the newly synthesized Linezolid analogues preserve the cellular integrity and
metabolic viability of ARPE-19 cells indicating no in vitro cytotoxic affects. These
Linezolid analogues may represent a new therapeutic alternative for treatment of the
disease cause by H. nana. Further studies will be conducted to study the activity of these
compounds against other intestinal parasites.
5
. Acknowledgements
Authors thank to Consejo Nacional de Ciencia y Tecnología (CONACYT) for the financial
support for these research (Grant No. SALUD-2015-1-261324), Supramolecular
Chemistry Thematic Network (Grant No. 294810), NMR facilities (Grant INFR-2011-3-
1
73395), X ray facilities (Grant INFR-2014-224405) and the scholarship program, which
supported to Eleazar Alcántar-Zavala and Esteban Hernández-Guevara for their
postgraduate studies. The docking software MOE was support by the Instituto de
Investigaciones Biomédicas at the UNAM, through the program “Novel alternatives for
the Treatment of Infection Deseases (NUATEI-IIB-UNAM)” and the program “Apoyo a
la Investigación y el Posgrado (PAIP)” Grant 5000-9163 in the Chemistry Faculty of the
UNAM.
2
1

6
. Supplementary information
FTIR, H and 13C NMR spectra are provided in a Supplementary Material File. All
crystallographic were deposited in the Cambridge Crystallographic Data Centre and the
accession numbers are 1958671, 1960623, 1960624 and 1960627.
1
7
. References
1
.- Langbang, D.; Dhodapkar, R.; Parija, S. C.; Premarajan, K. C. Rajkumari, N.
Prevalence of intestinal parasites among rural and urban population in Puducherry, South
India - A community-based study. J. Family Med. Prim. Care 2019, 8, 1607-1612.
doi:10.4103/jfmpc.jfmpc_196_19
2
.- Yang, D.; Zhao, W.; Zhang, Y.; Liu A. Prevalence of Hymenolepis nana and H.
diminuta from Brown Rats (Rattus norvegicus) in Heilongjiang Province, China. Korean
J. Parasitol. 2017, 55, 351-355. doi:10.3347/kjp.2017.55.3.351
3
.- Shahnazi, M.; Mehrizi, M. Z.; Alizadeh, S. A.; Heydarian, P.; Saraei, M.; Alipour, M.;
Hajialilo, E. Molecular characterization of Hymenolepis nana based on nuclear rDNA
ITS2 gene marker. Afr. Health Sci. 2019, 1, 1346-1352. doi:10.4314/ahs.v19i1.6
4
.- López-Angulo, G.; Verdugo-Gaxiola, S. E.; Montes-Avila, J.; Díaz-Camacho, S. P.;
Miranda-Soto, V.; Salazar-Salas, N. Y.; Delgado-Vargas, F. Bioguided isolation of N-
malonyl-(+)-tryptophan from the fruit of Pithecellobium dulce (Roxb.) Benth. that showed
high activity against Hymenolepis nana. Nat. Prod. Res. 2019, 33, 1-7.
doi:10.1080/14786419.2019.1590709
5
. Kandi V.; Koka S. S.; Bhoomigari, M. R. Hymenolepiasis in a Pregnant Woman: A
Case Report of Hymenolepis nana Infection. Cureus 2019, 11, e3810.
doi:10.7759/cureus.381
6
. Ahmed, O.; Elmesallamy, Gel-S.; Eassa, S. Comparative study of the hepatotoxic,
genotoxic and carcinogenic effects of praziquantel distocide and the natural myrrh extract
Mirazid on adult male albino rats. J. Egypt. Soc. Parasitol. 2005. 35, 313-29.
7
. Doenhoff, M. J.; Cioli, D.; Utzinger, J. Praziquantel: mechanisms of action, resistance
and new derivatives for schistosomiasis. Current Opinion in Infectious Diseases 2008, 21,
59-667. doi:10.1097/QCO.0b013e328318978f
6
2
2

8
. Jeziorski, M. C.; Greenberg, R. M. Voltage-gated calcium channel subunits from
platyhelminths: Potential role of praziquantel action. Int. J. Parasitol. 2006, 36, 625-636.
doi:10.1016/j.ijpara.2006.02.002.
9
. Ford, C. W.; Zurenko, G. E.; Barbachyn, M. R. The discovery of Linezolid, the first
oxazolidinone antibacterial agent. Curr. Drug Targets Infect. Disord. 2001, 2, 181-199.
doi:10.2174/1568005014606099
1
0. Hashemian, S. M. R.; Farhadi, T.; Ganjparvar, M. Linezolid: a review of its
properties, function, and use in critical care. Drug Des. Dev. Ther. 2018, 12, 1759-1767.
doi:10.2147/dddt.s164515
1
1. Saager, B.; Rohde, H.; Timmerbeil, B. S. Molecular characterization of Linezolid
resistance in two vancomycin-resistant (VanB) Enterococcus faecium isolates using
Pyrosequencing. Eur. J. Clin. Microbiol. Infect. Dis. 2008, 27, 873-878.
1
2. Besier, S.; Ludwig, A.; Zander, J.; Brade, V.; Wichelhaus, T. A. Linezolid resistance
in Staphylococcus aureus: gene dosage effect, stability, fitness costs, and cross-resistances.
Antimicrob. Agents Chem. 2008, 52, 1570-1572. doi:10.1128/aac.01098-07
1
3. Walked, J. A.; ODonnell, R.; Wiener, R. S. Linezolid vs glycopeptide antibiotics for
the treatment of suspected methicillin-resistant Staphylococcus aureus nosocomial
pneumonia: a meta-analysis of randomized controlled trials. Chest 2011, 139, 1148-1155.
doi:10.1016/j.yccm.2012.01.031
1
4. Zhao H.; Lu Y.; Sheng L.; Yuan Z.; Wang B.; Wang W.; Li Y.; Ma C.; Wang X.; Zhang
D.; Huang H. Discovery of fluorine-containing benzoxazinyl-oxazolidinones for the
treatment of Multidrug Resistant Tuberculosis. ACS Med. Chem. Lett. 2017, 8, 533−537.
doi:10.1021/acsmedchemlett.7b00068
1
5. Leach. K. L.; Swaney, S. M.; Colca, J. R. McDonald, W. G.; Blinn, J. R.; Thomasco,
L. M.; Gadwood, R. C.; Shinabarger, D.; Xion, L; Mankin, A. S. The site of action of
oxazolidinone antibiotics in living bacteria and in human mitochondria. Mol. Cell. 2007,
2
6, 393-402. doi:10.1016/j.molcel.2007.04.005
1
6. Ippolito, A. J.; Kanyo, F. Z.; Wang, D.; Franceschi, J. F.; Moore, B. P.; Steitz, A. T;
Duffy, M. E. Crystal structure of the oxazolidinone antibiotic Linezolid bound to the 50S
ribosomal subunit. J. Med. Chem. 2008, 51, 3353-3356. doi:10.1021/jm800379d
1
7. Ferrazzano, L.; Viola, A.; Lonati, E.; Bulbarelli, A.; Musumeci, R.; Cocuzza, C.;
Lombardo, M.; Tolomelli, A. New isoxazolidinone and 3,4-dehydro-b-proline derivatives
2
3

as antibacterial agents and MAO-inhibitors: A complex balance between two activities.
Eur. J. Med. Chem. 2016, 124, 906-919. doi:10.1016/j.ejmech.2016.09.007
1
8. Zhang, H.-Z.; Zhao, Z.-L.; Zhou, C.-H. Recent advance in oxazole-based medicinal
chemistry. Eur. J. Med. Chem. 2018, 144, 444–492. doi:10.1016/j.ejmech.2017.12.0
1
9. Barthel, D.; Schlitzer, M.; Pradel, G. Telithromycin and Quinupristin-Dalfopristin
induce delayed death in plasmodium falciparum. Antimicrob. Agents Chemother. 2008, 52,
7
2
74-777. doi:10.1128/aac.00892-07
0. Limoncu, M. E.; Eraç, B.; Gürpinar, T.; Özbilgin, A.; Balcıoğlu, İ. C.; Hoşgör-Limoncu
M. Investigation of in vitro antileishmanial activity of Moxifloxacin, Linezolid and
Caspofungin on Leishmania tropica promastigotes. Turkiye Parazitol Derg. 2013, 37, 1-3.
doi:10.5152/tpd.2013.01
2
1. Córdova-Guerrero, J.; Hernández-Guevara, E.; Ramírez-Zataraín, S.; Núñez-Bautista,
M.; Ochoa-Terán, A.; Muñiz-Salazar, R.; Montes-Ávila, J.; López-Angulo, G.; Paniagua-
Michel, A.; Nuño Torres, G. A. Antibacterial activity of new oxazolidin-2-one analogues
in methicillin-resistant Staphylococcus aureus strains. Int. J. Mol. Sci. 2014, 15, 5277-
5
291. doi:10.3390/ijms15045277
2
2. Ochoa-Terán, A.; Rivero, I. Synthesis of enantiopure 1, 3-oxazolidin-2-ones from α-
dibenzilamino esters. ARKIVOC 2008, 14, 330-343.18.
doi:http://dx.doi.org/10.3998/ark.5550190.0009.e30
2
3. Steven, J.; Brickner, S.; Hutchinson, D.; Barbachyn, M.; Mannimen, P.; Ulanowicz,
D.; Garmon, S.; Grega, K.; Hendges, S.; Toops, D.; Ford, C.; Zurenko, G. Synthesis and
antibacterial activity of U-100592 and U-100766, two oxazolidinone antibacterial agents
for the potential treatment of multidrug-resistant gram-positive bacterial infections. J. Med.
Chem. 1996, 39, 673-679.
2
4. Morán-Ramallal, R.; Liz, R.; and Gotor, V. Regioselective and stereospecific synthesis
of enantiopure 1,3-oxazolidin-2-ones by intramolecular ring opening of 2-(Boc-
aminomethyl)aziridines. Preparation of the antibiotic Linezolid. Org. Lett. 2008, 10, 1935-
1
938. doi:10.1021/ol800443p
2
5. Tohma, H.; Köksal, E.; Kilic, Ö.; Alan, Y.; Yilmaz, M.; Gülcin, I.; Bursal, E.; Alwasel,
S. RP-HPLC/MS/MS Analysis of the phenolic compounds, antioxidant and antimicrobial
activities of Salvia L. Species. Antioxidants 2016, 5, 38. doi:10.3390/antiox5040038
2
4

2
6. Gülçin, I.; Kirecci, E.; Akkemik, E.; Topal, F.; Hisar, O. Turk. Antioxidant,
antibacterial, and anticandidal activities of an aquatic plant: duckweed (Lemna minor L.
Lemnaceae). J. Biol. 2010, 34, 175-188. doi:10.1016/j.bioorg.2016.12.001
2
7. Kocyigit, U.; Budak, Y.; Gürdere, M.; Tekin, S.; Köprülü, T.; Ertürk, F.; Özcan, K.;
Gülçin, I.; Ceylan, M. Synthesis, characterization, anticancer, antimicrobial and carbonic
anhydrase inhibition profiles of novel (3aR,4S,7R,7aS)-2-(4-((E)-3-(3-aryl)acryloyl)
phenyl)-3a,4,7,7a-tetrahydro-1H-4,7-methanoisoindole-1,3(2H)-dione
Bioorg. Chem. 2017, 70,118-125. doi:10.1016/j.bioorg.2016.12.001.
derivatives.
2
8. Montes-Avila, J.; Díaz-Camacho, S. P.; Willms, K.; De la Cruz-Otero, M. C.; Robert,
L. Rivero, I. A.; Delgado-Vargas, F. Bioguided study of the in vitro parasitocidal effect on
adult Hymenolepis nana of the Psidium sartorianum (O. Berg Nied.) fruit methanol extract.
Med. Chem. Res. 2017, 26, 2845-2852. doi:10.1007/s00044-017-1983-x
2
9. Mosmann, T. Rapid colorimetric assay for cellular growth and survival: application to
proliferation and cytotoxicity assays. J. Immunol. Methods 1983, 65, 55-63.
doi:10.1016/0022-1759(83)90303-4
3
0. Hernandez, J.; Goycoolea, F.; Quintero, J.; Acosta, A.; Castañeda, M.; Dominguez, Z.;
Robles, R.; Vazquez-Moreno, L.; Velazquez, E.; Astiazaran, H.; Lugo, E.; Velazquez, C.
Sonoran propolis: chemical composition and antiproliferative activity on cancer cell lines.
Planta med. 2007. 73, 1469-1474. doi:10.1055/s-2007-990244
3
1. Desiraju, G. R.; Stainer, T. The weak hydrogen bond in structural chemistry and
biology. Oxford Universtiy Press, England. 1999, p. 13.
3
2. Molecular Operating Environment (MOE), version 2018.01, Chemical Computing
Group Inc., Montreal, Quebec, Canada. Available in http://www.chemcomp.com
3
3. García-Olaiz, G. D.; Alcántar-Zavala, E.; Ochoa-Terán, A.; Cabrera, A.; Muñiz-
Salazar, R.; Montes-Ávila, J.; Salazar-Medina, A. J.; Alday-Noriega, E.; Velazquez, C.;
Medina-Franco, J. L.; Laniado-Laborín, R. Design, synthesis and evaluation of the
antibacterial activity of new Linezolid dipeptide-type analogues. Bioorg. Chem. 2020, 95,
1
03483. doi:10.1016/j.bioorg.2019.103483
3
4. Schuwirth, B. S.; Borovinskaya, M. A.; Hau, C. W.; Zhang, W.; Vila-Sanjurjo,
A.; Holton, J. M. Crystal structure of the bacterial ribosome from Escherichia coli at 3.5
A resolution. Science 2005, 310, 827-834. doi:10.1126/science.1117230
2
5

3
5. Shaw, K. J.; Poppe, S.; Schaadt, R.; Brown-Driver, V.; Finn, J.; Pillar, C. M.;
Shinabarger, D.; Zurenko, G. In Vitro activity of TR-700, the antibacterial moiety of the
produg TR-701, against Linezolid-resistant strains. Antimicrob. Agents. Chemother. 2008,
5
2, 4442-4447. doi:10.1128/AAC.00859-08
3
6. Baily, G.; Garcia, H. H. Chapter 57, Other cestode infections: intestinal cestodes,
cysticercosis, other larval cestode infections. In: Manson’s Tropical Infectious Diseases,
2
014, 23 Ed, London UK: W. B. Saunders; p. 820-832.
3
7. Lipinski, C. A.; Lombardo, F.; Dominy, B. W.; Feeney, P. J. Experimental and
computational approaches to estimate solubility and permeability in drug discovery and
development settings. Adv. Drug Deliv. Rev. 2001, 46, 3-26. doi:10.1016/S0169-
4
09X(00)00129-0.
3
8. Lipinski, C. A. Lead- and drug-like compounds: the rule-of-five revolution. Drug
Discovery Today: Technologies 2004, 1, 337-341. doi:10.1016/j.ddtec.2004.11.007
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.
☐
The authors declare the following financial interests/personal relationships which may be
considered as potential competing interests:
2
6

Highlights

Stereoselective synthesis and characterization of six new Linezolid type
compounds.


The first Linezolid type compounds reported with in vitro antiparasitic activity
against Hymenolepis nana.
Promising potential as antiparasitic agents because they are more active and induce
anatomic damages not observed with Praziquantel.

These newly Linezolid analogues preserve the cellular integrity and metabolic
viability of ARPE-19 cells indicating no in vitro cytotoxic affects.
2
7