Rational design and synthesis of novel thiazolidin-4-ones as non-nucleoside HIV-1 reverse transcriptase inhibitors

Article abstract of DOI:10.1016/j.bmc.2014.04.018

A series of novel thiazolidin-4-one analogues, characterized by different substitution patterns at positions C-2 and N-3 of the thiazolidin-4-one scaffold for anti-HIV-1 activity has been investigated. Most of the compounds showed anti-HIV-1 activity at m

Full text of DOI:10.1016/j.bmc.2014.04.018

Accepted Manuscript
Rational design and synthesis of novel thiazolidin-4-ones as non-nucleoside
HIV-1 reverse transcriptase inhibitors
Vanangamudi Murugesan, Nandini Makwana, Rahul Suryawanshi, Reshu
Saxena, Rajkamal Tripathi, Ramesh Paranjape, Smita Kulkarni, Seturam B.
Katti
PII:
S0968-0896(14)00272-7
DOI:
http://dx.doi.org/10.1016/j.bmc.2014.04.018
BMC 11513
Reference:
Bioorganic & Medicinal Chemistry
To appear in:
Received Date:
Revised Date:
Accepted Date:
30 January 2014
9 April 2014
10 April 2014
Please cite this article as: Murugesan, V., Makwana, N., Suryawanshi, R., Saxena, R., Tripathi, R., Paranjape, R.,
Kulkarni, S., Katti, S.B., Rational design and synthesis of novel thiazolidin-4-ones as non-nucleoside HIV-1 reverse
transcriptase inhibitors, Bioorganic & Medicinal Chemistry (2014), doi: http://dx.doi.org/10.1016/j.bmc.
2014.04.018
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Rational design and synthesis of novel thiazolidin-4-ones as non-nucleoside
HIV-1 reverse transcriptase inhibitors
Vanangamudi Murugesana, Nandini Makwanac, Rahul Suryawanshic, Reshu Saxenab,
Rajkamal Tripathib, Ramesh Paranjapec, Smita Kulkarnic,$, Seturam B. Kattia,$,*
a Medicinal and Process Chemistry Division, CSIR-Central Drug Research Institute,
JankiPuram Extension, Sector-10, Lucknow-226031, Uttar Pradesh, India
b Toxicology Division, CSIR-Central Drug Research Institute (CDRI), JankiPuram Extension,
Sector-10, Lucknow-226031, Uttar Pradesh, India
c Department of Molecular Virology, National AIDS Research Institute (NARI), Pune-411026,
Maharashtra, India
$Equal contribution
*Corresponding author:
Dr. Seturam B. Katti, CSIR-CDRI, Lucknow, India
sb_katti@cdri.res.in
Tel: 0522-2772454
Fax: 0522-2771941
1

Abstract
A series of novel thiazolidin-4-one analogues, characterized by different substitution patterns
at positions C-2 and N-3 of the thiazolidin-4-one scaffold for anti-HIV-1 activity has been
investigated. Most of the compounds showed anti-HIV-1 activity at micromolar
concentrations when tested in TZM-bl cells in vitro. Among the thirty-three compounds
tested, compound 16 was the most potent inhibitor of HIV-1 replication against HIV-1IIIB,
HIV-1ADA5, HIV-1UG070 and HIV-1VB59 (EC50 = 0.02, 0.08, 0.08 and 0.08 µM, respectively)
with selectivity index (SI = 6940, 1735, 1692 and 1692) against tested viral strains,
respectively. The results of the present study suggested that the substitution of the nitro group
at 6' position of the C-2 phenyl ring and 2'',6''-dimethylpyridin-2-yl at the N-3 position of
thiazolidin-4-one had a major impact on the anti-HIV-1 activity and was found to lower
cytotoxicity. The substitution of the heteroaryl ring with bromo group and bicyclic heteroaryl
ring at N-3 thiazolidin-4-one was found to lower anti-HIV-1 activity and increase cytotoxicity.
The undertaken docking studies thus facilitated the identification of crucial interactions
between the HIV-1 RT enzyme and thiazolidin-4-one inhibitors, which can be used to design
new potential inhibitors.
Keywords: Thiazolidin-4-ones; Anti-HIV-1 activity; HIV-1 reverse transcriptase; NNRTIs
2

1. Introduction
The Non-nucleoside reverse transcriptase inhibitors (NNRTIs) play a vital role in the
management of HIV infection and are important component of highly active antiretroviral
therapy (HAART). They bind allosterically to the hydrophobic pocket located approximately
10 Å from the catalytic site in the palm domain of the p66 subunit, which harbor active sites
of the reverse transcriptase (RT) enzyme.1 Currently, the NNRTI class contains five FDA
approved drugs (nevirapine, delavirdine, efavirenz, etravirine, and rilpivirine) that are being
used in the management of HIV infection.2,3 However, these drugs have some common
adverse effects which include severe skin reactions, liver toxicity and sleep disorders. The
other complication that arises with this class of anti-retroviral drug is the emergence of drug
resistance due to rapid mutations in the amino acid residues of the NNRTI binding site.4-6
Hence, more efforts are needed in developing new anti-HIV-1 agents with higher potency that
would help in eradicating the emergence of pan-resistant viral variants. Among the five drugs,
etravirine and rilpivirine have been recognized as the most successful NNRTIs developed so
far due to their increased potency against HIV-1 wild-type (WT) and the clinically relevant
mutant strains.7 As reported in the earlier paper, substituted pyrimidin-2-yl and pyridin-2-yl
rings at the N-3 of thiazolidin-4-one derivatives were effective anti-HIV-1 agents with
minimal cytotoxicity.8,9 Recently, we described a new thiazolidin-4-one series based on
rational drug design approach in which the 5-methylisoxazol-3-yl moiety at N-3 of
thiazolidin-4-one showed inhibitory activity against HIV-1 reverse transcriptase.10 Moreover,
structural analysis of drugs/compound (rilpivirine, nevirapine and compound N-(4-bromo-1H-
pyrazol-3-yl)-2,1,3-benzothiadiazole-4-sulfonamide) demonstrated that fragments such as
para-cyanoaniline, pyridyl amide moiety (wing II of nevirapine) and an aryl sulfonamide
3

group, respectively seem to be essential moieties for anti-HIV-1 activity since they form
hydrophobic and hydrogen bonds interactions with the HIV-1 RT enzyme.11-13 Further, 3D-
QSAR of CoMFA and CoMSIA investigations performed on thiazolidin-4-one derivatives,
also suggested that the presence of steric groups at 2' and 6' positions of phenyl ring at C-2 of
thiazolidinone system and bulky groups at N-3 of heteroaromatic ring would influence
positively the HIV-1 RT inhibitory activity.14 Based on these results, we designed new
thiazolidin-4-one analogues in which a para-cyanophenyl moiety at C-2 or a 4-
methylbenzenesulfonamide, isonicotinamide, para-cyanoaniline and pyridylamino portions at
N-3 position were installed (Fig.1). Furthermore halogen atoms at 2',6'-positions of phenyl
ring were replaced by a nitro or methoxy groups. Additionally, other heteroaryl moieties were
introduced at N-3 of thiazolidin-4-one system. Here we report the structural modifications at
C-2 and N-3 of thiazolidin-4-one (Fig. 2) which could offer a structurally novel scaffold for
further development of a new series of thiazolidin-4-one as NNRTIs.
2. Results and discussion
2.1. Chemistry
The thiazolidin-4-ones (compounds 1-27) were synthesized by the multicomponent, one-pot
reaction conditions with the appropriate amino component and aldehyde in the presence of
mercaptoacetic acid in toluene under reflux conditions (Scheme 1).10 Whereas the synthesis of
thiazolidin-4-ones (compounds 28-33) was carried out in two steps. In the first step
hydrazones were synthesized by reacting aryl/heteroarylhydrazines and aromatic aldehydes in
toluene under reflux protocol and the toluene was removed, in the second step reaction
4

mixture was heated with large excess of mercaptoacetic acid under neat conditions at 60 °C
for 12 hours (Scheme 2).15
2.2. Anti-HIV-1 evaluation
The C-2 and N-3 modified thiazolidin-4-one compounds were tested against laboratory-
adapted HIV-1IIIB, HIV-1ADA5 strains and primary isolates (HIV-1UG070 and HIV-1VB59) in
TZM-bl cells using nevirapine as the reference molecule. The results of anti-HIV-1 activity,
cytotoxicity and selectivity index of thiazolidin-4-ones were expressed as EC50, CC50, and SI
values, respectively (Table 1 and Table 2). In addition, these compounds were tested for
inhibition of HIV-1 reverse transcriptase enzyme, using a RT assay kit, and expressed as the
percentage inhibitions at 100 µg/ml (Table 1). The potency of the some thiazolidin-4-ones
displayed moderate to potent activities against the four tested HIV-1 strains with EC50 values
in the range of 11.82–0.02 µM concentrations. Among all the thiazolidin-4-ones, compound
16 was the most potent inhibitor of HIV-1 replication against HIV-1IIIB, HIV-1ADA5, HIV-
1UG070 and HIV-1VB59 (EC50 = 0.02, 0.08, 0.08 and 0.08 µM, respectively), with the highest
selectivity index (SI = 6940, 1735, 1692 and 1692, respectively). Compound 16 exhibited
similar HIV-1IIIB inhibitory activity, when compared with the reference compound nevirapine
(EC50 = 0.03 µM). But in other primary isolates (HIV-1UG070 and HIV-1VB59), compound 16
(EC50 = 0.08 µM) showed 2.5 less fold activity than the positive control nevirapine. In
addition, the compound 16 exhibited less cytotoxicity (CC50 = 138.8 µM in TZM-bl cells) and
also showed 91.55% inhibition against HIV-1 RT enzyme. Taking into consideration the
substitution at N-3, from the data reported in Table 1 it can be observed that the compounds
which present a 4,6-dimethyl pyiridin-2-yl substitution at N-3 show a better antiretroviral
activity profile than the corresponding compounds with other heteroaryl moieties (see
5

compounds 3 and 16). The 4-6-dimethoxypyrimidin-2-yl substitution also seems to have
positive effects on the anti HIV-1 activity (see compounds 17 and 25-27) and in particular on
% RT inhibition of compounds 21 and 26 which is 93. 81% and 95.40%, respectively.
Furthermore, the presence of bromine atom at 5 position on the 4,6-dimethylpyiridin-2-yl
nucleus at N-3 position marginally reduced the potency and subsequently increase the cellular
toxicity profile of compound 19 and the previously reported corresponding 5-bromo-4,6-di-
me-pyridin-2-yl congeners.10 Introduction of a bicyclic ring of 9H-purin-6-yl at N-3 of
thiazolidin-4-one in compounds 23 and 24 (EC50 = 11.82 and 36.33 µM against HIV-1IIIB and
11.71 and 27.84 µM against HIV-1ADA5 strains, respectively) led to reduced anti-viral potency.
These observations suggested that the magnitude of the bicyclic ring did not accommodate in
the binding pocket of RT. In the set of 4,6-di-methoxypyrimidin-2-yl congeners at N-3,
compounds 17, 26 and 27 revealed slight reduction of anti-HIV-1 activities than compound 25
because of replacement of the Cl atom with -NO2 group or F atom at 6' position of the C-2
phenyl ring of thiazolidin-4-one. Based on the results it may be inferred that introduction of -
NO2 group at 6' position of the C-2 phenyl ring of thiazolidin-4-one has a positive influence
on the anti-HIV-1 activity (when 4,6-di-Me-pyridin-2-yl moiety placed at N-3). Moreover the
compounds 20-22, having 2’,6’-dimethoxyphenyl moiety at C-2 of thiazolidin-4-one,
displayed moderate to poor activity against all four HIV-1 strains. The introduction of a para-
cyanoaniline in compounds 1-8 in place of 2',6'-dihalophenyl group at C-2 of thiazolidin-4-
one, leads to a substantial decrease of potency in both enzyme inhibition and anti-viral
activity in cell cultures. Additionally, the impact of introducing 3-cyanopyridin-6-yl moiety in
compound 9 at the N-3 of thiazolidin-4-one, resulted in low or moderate activities against the
four HIV-1 strains and loss of HIV-1 RT inhibition. Further, we evaluated the effect of a
6

linker (-NH-, -CONH-, and -SO2NH-) between heteroaryl/aryl rings and the N-3 position of
thiazolidin-4-one on anti-HIV-1 activity. Only the NH- linker of pyridyl-2-ylamino moiety led
to compounds 28 and 29 which showed good to moderate potency with EC50 = 0.99 and 2.10
µM against HIV-1IIIB and 1.85 and 3.79 µM against HIV-1ADA5, 1.17 and 4.66 µM against
HIV-1UGO70 and 1.70 and 5.31 µM against HIV-1VB59 strains respectively. The other linkers
mainly -CONH- and -SO2NH- present in compounds 10-15 showed very low potency against
all HIV-1 strains with concomitant decrease of the RT inhibition. This indicated that the linker
between heteroaryl/aryl rings and N-3 of thiazolidin-4-one core could play a crucial role in the
antiviral activity and HIV-1 RT enzyme inhibition.
2.3. Molecular Docking Studies
The molecular interactions studies of newly synthesized thiazolidin-4-ones were explored by
using the AutoDock 4.016. Five compounds, 10, 12, 16, 25 and 28, were docked into the active
site of HIV-1 RT non-nucleoside binding site (NNBS). The coordinates of the non-nucleoside
binding site were taken from the crystal structure of HIV-1 Reverse Transcriptase (RT) in
complex with TMC278 (Rilpivirine) (pdb entry codes: 2ZD1)17. Detailed analysis of the
binding mode of potent compound 16 (Fig. 3) showed that the 4,6-dimethylpyridin-2-yl
moiety along with thiazolidin-4-one scaffold lay in a hydrophobic pocket, which is mainly
formed by the side chains of Tyr181, Tyr188, Phe227, and Trp229. The nitro group at 6'-
phenyl moiety in compound 16 also formed two hydrogen bonds with the backbone of Gly190
and Val189 in the HIV-1 RT. The 2',6'-dihalophenyl moiety along with thiazolidin-4-one ring
of compound 25 and 2',6'-dihalophenyl part of compounds 10, 12 and 28 (Fig. 4 and 5) exist
in the π-box formed by the hydrophobic residues of Tyr181, Tyr188, Phe227, and Trp229.
The oxygen atom of the terminal dimethoxy group at pyrimidin-2-yl of compound 25 formed
7

two hydrogen bonds with the backbone amino groups of Lys101 and Lys103 residues. The
carbonyl oxygen atom of the thiazolidin-4-one nucleus and -NH-SO2- group of compounds 12
and 10 forms single hydrogen bond interactions with the backbone N–H of Lys101 residue. In
the compound 28, the pyridyl-2yl ring makes close hydrophobic contacts with the aliphatic
isobutyl side chain of Leu100 and Leu234. The docked binding energy (Table 3), of
compounds 10, 12, 16, 25 and 28 displayed moderate to strong binding energy values (binding
energy = -8.23, -7.98, -10.08, -8.96 and -9.67 kcal/mol, respectively). The thiazolidin-4-ones
(16 and 28) form stable complexes with the RT enzyme, because of various hydrogen bonds
and hydrophobic interactions with the RT enzyme. Molecular modeling results could be used
as a support for the development of novel and more active thiazolidin-4-ones against the HIV-
1 RT target.
3. Conclusions
In this study, we have discussed the preliminary structure-activity relationships of C-2 and N-
3 modified thiazolidin-4-one derivatives against four HIV-1IIIB, HIV-1ADA5, HIV-1UG070 and
HIV-1VB59 strains along with HIV-1 RT enzyme assay. Amongst these, compound 16 was
found to be the most promising inhibitor of HIV-1 replication against HIV-1IIIB, HIV-1ADA5
,
HIV-1UG070 and HIV-1VB59 (EC50 = 0.02, 0.08, 0.08 and 0.08 µM, respectively), with highest
selectivity index SI = 6940, 1735, 1692 and 1692, respectively. The overall SAR information
and binding model analysis of thiazolidin-4-one analogues could serve as potential lead to
further development of novel and potent NNRTIs.
8

4. Experimental protocol
4.1. General
All chemicals and solvents were purchased from Sigma Aldrich and Merck, respectively.
Melting points (Mp) were determined by a Complab apparatus and are uncorrected. Infrared
(IR) spectra were recorded with a Perkin Elmer Spectrum RX1 spectrometer in potassium
bromide (4000-450 cm-1). 1H-NMR and 13C-NMR spectra were recorded on a NMR
Spectrometer Bruker DRX-300 using CDCl3 as a solvent at 300 and 75 MHz, respectively.
The chemical shifts are reported as parts per million (δ ppm) from tetramethylsilane (TMS) as
an internal standard. The following abbreviations are used to describe peak patterns: s
(singlet), d (doublet), dd (double doublet), t (triplet), m (multiplet), q (quadruplet), and br
(broad). Coupling constants are given in hertz. HRMS and ESI-MS were performed using a
Waters Agilent 6520- Q-TofMS/MS system and JEOL-AccuTOF JMST100LC instruments.
Elemental analyses were carried out on CARLO-ERBA EA1108 C, H, N elemental analyzer
and values were in the acceptable limits of the calculated values. Analytical thin-layer
chromatography (TLC) was carried out on Merck’s precoated silica-gel plates 60 F254 and
spots were visualized by irradiation with UV light (254 nm) and/or by iodine vapours. Column
chromatography separations were performed on silica gel (230-400 mesh) as the stationary
phase using hexane-ethyl acetate as mobile phase.
4.2. General synthetic procedure for the compounds from (hetero) aromatic
amine/tosylhydrazine/isonicotinohydrazide (compounds 1-27)
The appropriate (hetero) aromatic amine/tosylhydrazine/isonicotinohydrazide (1.0 mmol) and
2,6-disubstituted benzaldehyde/4-cyanobenzaldehyde (1.5 mmol) were stirred in dry toluene
9

under reflux condition followed by addition of mercaptoacetic acid (2.0 mmol). The reaction
mixture was refluxed with a Dean-Stark trap for 24-48 hours until the complete consumption
of (hetero) aromatic amine. The reaction mixture was concentrated to dryness under reduced
pressure and the residue was taken up in ethyl acetate. The organic layer was successively
washed with 5% aqueous citric acid, water, 5% aqueous sodium hydrogen carbonate, and then
finally with brine. The organic layer was dried over sodium sulfate and the solvent was
removed under reduced pressure to get a crude product that was purified by column
chromatography on silica-gel (230-400 mesh) using hexane–ethyl acetate as eluents. The
structures of all synthesized compounds were characterized by means of TLC, IR, ESI-MS, 1H
NMR, 13C NMR, elemental analyses and HRMS.
4.2.1. 4-(3-(5-methylisoxazol-3-yl)-4-oxothiazolidin-2-yl)benzonitrile (1)
White solid, yield: 98.7%. Mp: 95-96 °C; IR (KBr): νmax C=O 1708, CN 2229 cm-1; 1H NMR
(300 MHz, CDCl3); δ 7.66 (d, 2H, J = 8.2 Hz, H2,6-PhH), 7.41 (d, 2H, J = 8.2 Hz, H3,5-PhH),
6.84 (s, 1H, H4-isoxazol-3-yl-H), 6.34 (s, 1H, CH), 3.98 (d, 1H, J = 16.2 Hz, CH2), 3.79 (d,
1H, J = 16.3 Hz, CH2), 2.39 (s, 3H, CH3); 13C NMR (75 MHz, CDCl3): δ 170.6 (C=O), 170.5
(isoxazol-3-yl-C5), 157.2 (isoxazol-3-yl-C3), 145.6, 132.8, 126.3, (Ph-C), 118.3 (CN), 112.3
(Ph-C), 95.4 (isoxazol-3-yl-C4), 61.3 (C2), 34.3 (C5), 12.6 (CH3); MS (ESI): m/z 285.1 (M+).
Anal. Calcd for C14H11N3O2S: C, 58.93; H, 3.89; N, 14.73. Found: C, 58.90; H, 3.91; N,
14.77.
4.2.2. 4-(3-(5-bromo-4,6-dimethylpyridin-2-yl)-4-oxothiazolidin-2-yl)benzonitrile (2)
White solid, yield: 94.6 %. Mp: 155-156 °C; IR (KBr): νmax C=O 1710, CN 2222 cm-1; 1H
NMR (300 MHz, CDCl3); δ 7.93 (s, 1H, H3-pyridin-2-yl), 7.61 (d, 2H, J = 8.3 Hz, H2,6-PhH),
7.42 (d, 2H, J = 8.2 Hz, H3,5-PhH), 6.81 (s, 1H, CH), 4.05 (dd, 1H, J = 16.1 Hz, 0.9 Hz, CH2),
10

3.86 (d, 1H, J = 16.1 Hz, CH2), 2.41 (d, 6H, J = 1.0 Hz, CH3 at C4,6-pyridin-2-yl); 13C NMR
(75 MHz, CDCl3): δ 171.7 (C=O), 155.3, (pyridin-2-yl-C6), 149.5 (pyridin-2-yl-C4), 147.7,
147.0 (pyridin-2-yl-C2 & Ph-C4), 132.5 (Ph-C2 & C6), 126.6, (Ph-C3 & C5), 120.0 (pyridin-
2-yl-C5),118.3 (CN), 115.4 (pyridin-2-yl-C3), 111.9 (Ph-C1), 62.2 (C2), 34.0 (C5), 25.0, 23.7
(CH3 at C4,6-pyridin-2-yl); MS (ESI): m/z 389.0 (M+H+). HRMS (ESI): (M+H+), calcd. for
C17H14BrN3OS+H+ m/z 388.0119, Found 388.0122.
4.2.3. 4-(3-(4,6-dimethylpyridin-2-yl)-4-oxothiazolidin-2-yl)benzonitrile (3)
White solid, yield: 95.8 %. Mp: 145-146 °C; IR (KBr): νmax C=O 1690, CN 2228 cm-1; 1H
NMR (300 MHz, CDCl3); δ 7.75 (s, 1H, H3-pyridin-2-yl), 7.59 (d, 2H, J = 8.2 Hz, H2,6-PhH),
7.43 (d, 2H, J = 8.2 Hz, H3,5-PhH), 6.88 (s, 1H, H5-pyridin-2-yl), 6.71 (s, 1H, CH), 4.05 (d,
1H, J = 16.0 Hz, CH2), 3.87 (d, 1H, J = 16.0 Hz, CH2), 2.32, 2.24 (s, 6H, CH3 at C4,6-pyridin-
2-yl); 13C NMR (75 MHz, CDCl3): δ 170.0 (C=O), 156.2 (pyridin-2-yl-C6), 149.6 (pyridin-2-
yl-C4), 147.1 (pyridin-2-yl-C2 & Ph-C4), 132.4 (Ph-C2 & C6), 126.8 (Ph-C3 & C5), 121.46
(pyridin-2-yl-C5), 118.4 (CN), 114.1 (pyridin-2-yl-C3), 111.6 (Ph-C1), 62.4 (C2), 34.1 (C5)
23.6, 21.2 (CH3 at C4,6-pyridin-2-yl); MS (ESI): m/z 310.0 (M+H+). HRMS (ESI): (M+H+),
calcd. for C17H15N3OS+H+ m/z 310.1014, Found 310.1045.
4.2.4. 4-(3-(4,6-dimethoxypyrimidin-2-yl)-4-oxothiazolidin-2-yl)benzonitrile (4)
White solid, yield: 93.1 %. Mp: 138-137 °C; IR (KBr): νmax C=O 1734, CN 2224 cm-1; 1H
NMR (300 MHz, CDCl3); δ 7.64 (d, 2H, J = 8.2 Hz, H2,6-PhH), 7.46 (d, 2H, J = 8.2 Hz, H3,5-
PhH), 6.57 (s, 1H, H5-pyrimidin-2-yl), 5.78 (s, 1H, CH), 3.99 (d, 2H, J = 16.1 Hz, CH2), 3.80
(s, 6H, OCH3 at C4,6-pyrimidin-2-yl); 13C NMR (75 MHz, CDCl3): δ 171.5 (C=O), 169.7
(pyrimidin-2-yl-C4 & C6), 155.1 (pyrimidin-2-yl-C2), 147.0 (Ph-C4), 132.7 (Ph-C2 & C6),
126.0 (Ph-C3 & C5), 118.2 (CN), 112.0 (Ph-C1), 86.7 (pyrimidin-2-yl-C5), 62.4 (C2), 54.2
11

(OCH3 at C4,6-pyrimidin-2-yl), 33.7 (C5); MS (ESI): m/z 343 (M+H+). HRMS (ESI): (M+H+),
calcd. for C16H14N4O3S+H+ m/z 343.0865, Found 343.0891.
4.2.5. 4-(3-(4,6-dimethylpyrimidin-2-yl)-4-oxothiazolidin-2-yl)benzonitrile (5)
Pale Yellow solid, yield: 84.9 %. Mp: 161-162 °C; IR (KBr): νmax C=O 1710, CN 2227 cm-1;
1H NMR (300 MHz, CDCl3); δ 7.59 (dd, 4H, J = 8.3 Hz, 13.9 Hz, H2,3,5,6-PhH), 6.78 (s, 1H,
H5-pyrimidin-2-yl), 6.69 (s, 1H, CH), 4.03 (dd, 1H, J = 16.1 Hz, 1.0 Hz, CH2), 3.89 (d, 1H, J
= 16.0 Hz, CH2), 2.39 (s, 6H, CH3 at C4,6-pyrimidin-2-yl); 13C NMR (75 MHz, CDCl3): δ
170.3 (C=O), 168.3 (pyrimidin-2-yl-C4 & C6), 156.2 (pyrimidin-2-yl-C2), 146.0 (Ph-C4),
132.4 (Ph-C2 & C6), 127.3 (Ph-C3 & C5), 118.3 (CN), 117.5 (pyrimidin-2-yl-C5), 112.1 (Ph-
C1), 62.6 (C2), 34.0 (C5), 23.8 (CH3 at C4,6-pyrimidin-2-yl); MS (ESI): m/z 311.0 (M+H+).
HRMS (ESI): (M+H+), calcd. for C16H14N4OS+H+ m/z 311.0967, Found 311.0978
4.2.6. 4-(3-(4-methylpyrimidin-2-yl)-4-oxothiazolidin-2-yl)benzonitrile (6)
Pale Yellow solid, yield: 86.8 %. Mp: 163-164 °C; IR (KBr): νmax C=O 1716, CN 2230 cm-1;
1H NMR (300 MHz, CDCl3); δ 8.49 (d, 1H, J = 5.0 Hz, H6-pyrimidin-2-yl), 7.60 (dd, 4H, J =
8.3 Hz, 23.3 Hz, H2,3,5,6-PhH), 6.92 (d, 1H, J = 5.0 Hz, H5-pyrimidin-2-yl), 6.69 (s, 1H, CH),
4.04 (d, 1H, J = 16.1 Hz, CH2), 3.90 (d, 1H, J = 16.1 Hz, CH2), 2.43 (s, 3H, CH3 at C4-
pyrimidin-2-yl); 13C NMR (75 MHz, CDCl3): δ 170.3 (C=O), 168.9 (pyrimidin-2-yl-C4),
157.8 (pyrimidin-2-yl-C2), 156.5 (pyrimidin-2-yl-C6), 146.1 (Ph-C4), 132.6 (Ph-C2 & C6),
127.1 (Ph-C3 & C5), 118.4 (CN), 117.9 (pyrimidin-2-yl-C5), 112.2 (Ph-C1), 62.6 (C2), 34.0
(C5), 24.6 (CH3 at C4-pyrimidin-2-yl); MS (ESI): m/z 297.0 (M+H+). Anal. Calcd for
C15H12N4OS: C, 60.79; H, 4.08; N, 18.91. Found: C, 60.72; H, 4.11; N, 18.88
12

4.2.7. 4,4'-(4-oxothiazolidine-2,3-diyl)dibenzonitrile (7)
Yellow solid, yield: 98.4 %. Mp: 152-153 °C; IR (KBr): νmax C=O 1703, CN 2223 cm-1; 1H
NMR (300 MHz, CDCl3); δ 7.65-7.58 (m, 3H, H2,3,6-Ph-4'-yl), 7.41 (d, 3H, J = 8.4 Hz, H2,3,6
Ph-4-yl), 7.14-7.03 (m, 1H, H5-Ph-4'-yl), 6.85 (d, 1H, J = 7.5 Hz, H5-Ph-4-yl), 6.22 (s, 1H,
CH), 4.00 (d, 1H, J = 16.0 Hz, CH2), 3.90 (d, 1H, J = 16.1 Hz, CH2); 13C NMR (75 MHz,
CDCl3): δ 170.8 (C=O), 143.9, 141.1, 133.1, 127.1, 124.2, (Ph-C & C'), 117.9, 117.9 (di-
-
CN), 113.1, 110.1 (Ph-C & C'), 63.7 (C2), 33.7 (C5); MS (ESI): m/z 305.2 (M+). Anal. Calcd
for C17H11N3OS: C, 66.87; H, 3.63; N, 13.76. Found: C, 66.90; H, 3.61; N, 13.74.
4.2.8. 6-(2-(4-cyanophenyl)-4-oxothiazolidin-3-yl)nicotinonitrile (8)
White solid, yield: 88.3 %. Mp: 129-130 °C; IR (KBr): νmax C=O 1708, CN 2231 cm-1; 1H
NMR (300 MHz, CDCl3); δ 8.55 (d, 1H, J = 8.8 Hz, H2-pyridin-6-yl), 8.47 (d, 1H, J = 1.6 Hz,
H4-pyridin-6-yl), 8.01 (dd, 1H, J = 8.8 Hz, 2.2 Hz, H5-pyridin-6-yl), 7.64 (d, 2H, J = 8.3 Hz,
H3,5-PhH), 7.39 (d, 2H, J = 8.2 Hz, H2,6-PhH), 6.81 (s, 1H, CH), 4.06 (d, 1H, J = 16.5 Hz,
CH2), 3.87 (d, 1H, J = 16.5 Hz, CH2); 13C NMR (75 MHz, CDCl3): δ 171.5 (C=O), 152.6
(pyridin-6-yl-C2), 150.9 (pyridin-6-yl-C6), 146.2 (Ph-C4), 141.2 (pyridin-6-yl-C4), 132.8
(Ph-C3 & C5), 126.2 (Ph-C2 & C6), 118.2, 116.2 (di-CN), 115.1 (pyridin-6-yl-C5), 112.2
(Ph-C1), 106.1 (pyridin-6-yl-C3) 62.0 (C2), 33.7 (C5); MS (ESI): m/z 306.2 (M+). Anal.
Calcd for C16H10N4OS: C, 62.73; H, 3.29; N, 18.29. Found: C, 62.78; H, 3.25; N, 18.34.
4.2.9. 6-(2-(2,6-dichlorophenyl)-4-oxothiazolidin-3-yl)nicotinonitrile (9)
White solid, yield: 85.9 %. Mp: 119-120 °C; IR (KBr): νmax C=O 1712, CN 2225 cm-1; 1H
NMR (300 MHz, CDCl3); δ 8.50-8.46 (m, 2H, H2,4-pyridin-6-yl), 7.96-7.92 (m, 1H, H5-
pyridin-6-yl), 7.54 (s, 1H, CH), 7.36-7.33 (m, 1H, H4-PhH), 7.19-7.09 (m, 2H, H3,5-PhH),
4.23 (dd, 1H, J = 16.1 Hz, 1.6 Hz, CH2), 3.98 (d, 1H, J = 16.1 Hz, CH2); 13C NMR (75 MHz,
13

CDCl3): δ 172.0 (C=O), 152.9 (pyridin-6-yl-C2), 150.6 (pyridin-6-yl-C6), 140.9 (pyridin-6-
yl-C4), 135.5 (Ph-C1), 134.2, 132.8 (Ph-C2 & C6), 130.5, 129.4 (Ph-C3 & C5), 128.8 (Ph-
C4), 116.4 (CN), 115.3 (pyridin-6-yl-C5), 105.8 (pyridin-6-yl-C3) 59.0 (C2), 35.5 (C5); MS
(ESI): m/z 350.0 (M+). Anal. Calcd for C15H9Cl2N3OS: C, 51.44; H, 2.59; N, 12.00. Found: C,
51.41; H, 2.60; N, 12.03.
4.2.10. N-(2-(2,6-dichlorophenyl)-4-oxothiazolidin-3-yl)-4-methylbenzenesulfonamide(10)
Pale yellow solid, yield: 89.1%. Mp: 140-142 °C; IR (KBr): νmax C=O 1699, -NH- 3399, -N-
SO2- 1327, 1220 cm-1; 1H NMR (300 MHz, CDCl3); δ 8.08, 7.99 (s, 2H, H2,6-Ph-SO2-NH-),
7.93 (d, 2H, J = 8.2 Hz, H3,5-Ph-SO2-NH-), 7.35 (t, 3H, J = 7.0 Hz, H3,4,5-PhH), 7.23 (s, 1H,
CH), 4.17 (d, 1H, J = 15.14 Hz, CH2), 4.13 (d, 1H, J = 15.0 Hz, CH2), 2.44 (s, 3H, CH3 at C4-
Ph-SO2-NH-); 13C NMR (75 MHz, CDCl3): δ 165.3 (C=O), 146.7, 142.9, 138.3, 137.3, 137.2,
133.6, 132.1, 130.4, 128.4, 123.1, 122.9, 117.7, (Ph-C), 51.9 (C2), 35.2 (C5), 23.7 (CH3); MS
(ESI): m/z 417.9 (M+). Anal. Calcd for C16H14Cl2N2O3S2: C, 46.05; H, 3.38; N, 6.71. Found:
C, 46.00; H, 3.32; N, 6.79.
4.2.11.N-(2-(2-chloro-6-fluorophenyl)-4-oxothiazolidin-3-yl)-4
methylbenzenesulfonamide (11)
Pale yellow solid, yield: 87.3%. Mp: 139-141 °C; IR (KBr): νmax C=O 1703, -NH- 3451, -N-
SO2- 1323, 1222 cm-1; 1H NMR (300 MHz, CDCl3); δ 8.04 (d, 2H, J = 7.1 Hz, H2,6-Ph-SO2-
NH), 7.93 (d, 2H, J = 8.2 Hz, H3,5-Ph-SO2-NH-), 7.36 (d, 3H, J = 7.9 Hz, H3,4,5-PhH), 7.03 (t,
1H, J = 8.2, CH), 4.17 (d, 1H, J = 15.1 Hz, CH2), 4.12 (d, 1H, J = 15.2 Hz, CH2), 2.44 (s, 3H,
CH3 at C4- Ph-SO2-NH-); 13C NMR (75 MHz, CDCl3): δ 165.8 (C=O), 162.4, 147.2, 140.6,
138.6, 134.4, 134.2, 132.6, 130.9, 115.1, 115.0, 114.8, 114.6, 52.9 (C2), 34.3 (C5), 24.6
14

(CH3); MS (ESI): m/z 400.4 (M+). Anal. Calcd for C16H14ClFN2O3S2: C, 47.94; H, 3.52; N,
6.99. Found: C, 47.98; H, 3.45; N, 6.82.
4.2.12. N-(2-(2,6-dichlorophenyl)-4-oxothiazolidin-3-yl)isonicotinamide (12)
Yellow solid, yield: 91.2%. Mp: 139-141 °C; IR (KBr): νmax -NH- 3342, thiazolidinone C=O
1719, amide C=O 1647 cm-1; 1H NMR (300 MHz, CDCl3); δ 8.47 (s, 2H, H2,6-
isonicotinamide), 8.39 (br, 1H, CONH), 7.39-7.35 (m, 2H, H3,5-isonicotinamide), 7.23-7.14
(m, 3H, H3,4,5-PhH), 7.06 (s, 1H, CH), 4.08 (dd, 1H, J = 15.6 Hz, 1.8 Hz, CH2), 3.91 (d, 1H, J
= 15.6 Hz, CH2); 13C NMR (75 MHz, CDCl3): δ 172.7 (thiazolidinone C=O), 167.9 (amide
C=O), 145.8, 135.1, 135.0, 134.6, 132.1, 131.0, 130.6, 130.4, 130.2, 128.8, 128.6 (aromatic-
C), 57.9 (C2), 32.3 (C5); MS (ESI): m/z 368.1 (M+). Anal. Calcd for C15H11Cl2N3O2S: C,
48.93; H, 3.01; N, 11.41. Found: C, 48.98; H, 3.09; N, 11.38.
4.2.13. N-(2-(2-chloro-6-fluorophenyl)-4-oxothiazolidin-3-yl)isonicotinamide (13)
Yellow solid, yield: 93.7%. Mp: 125-126 °C; IR (KBr): νmax -NH- 3348, thiazolidinone C=O
1709, amide C=O 1666 cm-1; 1H NMR (300 MHz, CDCl3); δ 8.98 (br, 1H, CONH), 8.74 (s,
1H, H2-isonicotinamide), 7.36-7.29 (m, 2H, H6,3-isonicotinamide), 6.98-6.85 (m, 4H, H5-
isonicotinamide & H3,4,5-PhH), 6.51 (s, 1H, CH), 4.11 (d, 1H, J = 15.5 Hz, CH2), 3.82 (d, 1H,
J = 15.6 Hz, CH2); 13C NMR (75 MHz, CDCl3): δ 172.7 (thiazolidinone C=O), 163.1 (amide
C=O), 159.7, 133.4, 131.3, 131.1, 126.0, 124.0, 123.8, 115.6, 115.3 (aromatic-C), 74.7 (C2),
32.0 (C5); MS (ESI): m/z 351.2 (M+). Anal. Calcd for C15H11ClFN3O2S: C, 51.21; H, 3.15; N,
11.94. Found: C, 51.29; H, 3.07; N, 11.97.
4.2.14. N-(2-(4-nitrophenyl)-4-oxothiazolidin-3-yl)isonicotinamide (14)
Yellow solid, yield: 90.8%. Mp: 136-137 °C; IR (KBr): νmax -NH- 3348, thiazolidinone C=O
1712, amide C=O 1656 cm-1; 1H NMR (300 MHz, CDCl3); δ 9.06 (s, 1H, CONH), 8.29 (d,
15

2H, J = 8.7 Hz, H2,6-isonicotinamide), 8.22 (d, 2H, J = 8.7 Hz, H3,5-PhH), 7.76 (d, 2H, J = 8.7
Hz, H3,5-isonicotinamide), 7.57 (d, 2H, J = 8.7 Hz, H2,6-PhH), 6.19 (s, 1H, CH), 3.95 (d, 1H, J
= 16.1 Hz, CH2), 3.86 (d, 1H, J = 16.1 Hz, CH2); 13C NMR (75 MHz, CDCl3): δ 168.0
(thiazolidinone C=O), 150.65 (amide C=O), 149.0, 148.1, 146.1, 139.4, 128.6, 128.3, 127.4,
125.1, 124.4, 123.9 (aromatic-C), 63.5 (C2), 31.3 (C5); MS (ESI): m/z 345.1 (M+H+). Anal.
Calcd for C15H12N4O4S: C, 52.32; H, 3.51; N, 16.27. Found: C, 52.25; H, 3.59; N, 16.31.
4.2.15. N-(2-(2,6-dimethylphenyl)-4-oxothiazolidin-3-yl)isonicotinamide (15)
White solid, yield: 88.3%. Mp: 155-157 °C; IR (KBr): νmax -NH- 3342, thiazolidinone C=O
1703, amide C=O 1626 cm-1; 1H NMR (300 MHz, CDCl3); δ 8.50 (s, 1H, CONH), 7.15-6.96
(m, 7H, H2,3,5,6-isonicotinamide & H3,4,5-PhH ), 6.70 (s, 1H, CH), 3.89 (s, 2H, CH2), 2.19 (s,
6H, di-CH3 at C2,6-Ph); 13C NMR (75 MHz, CDCl3): δ 167.0 (thiazolidinone C=O), 151.8
(amide C=O), 137.9, 137.2, 135.7, 132.4, 131.2, 131.1, 129.3, 129.1, 128.8, 128.5 (aromatic-
C), 59.1 (C2), 31.9 (C5), 20.6, 20.4 (CH3 at C2,6-Ph); MS (ESI): m/z 327.1 (M+). Anal. Calcd
for C17H17N3O2S: C, 62.36; H, 5.23; N, 12.83. Found: C, 62.31; H, 5.20; N, 12.88.
4.2.16. 2-(2-chloro-6-nitrophenyl)-3-(4,6-dimethylpyridin-2-yl)thiazolidin-4-one (16)
White solid, yield: 77.2%. Mp: 108-109 °C; IR (KBr): νmax C=O 1695cm-1; 1H NMR (300
MHz, CDCl3); δ 7.86 (s, 1H, H5-PhH), 7.65 (d, 1H, J = 6.3 Hz, H3-PhH), 7.48 (d, 1H, J = 7.6
Hz, H4-PhH), 7.30, 7.25 (s, 2H, H3,5-pyridin-2-yl), 6.64 (s, 1H, CH), 4.23 (d, 1H, J = 15.8 Hz,
CH2), 3.99 (d, 1H, J = 15.8 Hz, CH2), 2.28 (s, 3H, CH3 at C6-pyridin-2-yl), 2.16 (s, 3H, CH3 at
C4-pyridin-2-yl); 13C NMR (75 MHz, CDCl3): δ 171.1 (C=O), 156.1 (pyridin-2-yl-C6), 149.6,
149.49 (pyridin-2-yl-C4 & Ph-C6), 135.6 (pyridin-2-yl-C2), 133.8, 132.8. 128.6, 122.3, 121.2
(Ph-C), 112.8 (pyridin-2-yl-C3 & C5), 57.1 (C2). 35.9 (C5) 23.1, 21.2 (CH3 at C4,6-pyridin-2-
16

yl); MS (ESI): m/z 364.0 (M+H+). HRMS (ESI): (M+H+), calcd. for C16H14ClN3O3S+H+ m/z
364.0523, Found 364.0585.
4.2.17. 2-(2-chloro-6-nitrophenyl)-3-(4,6-dimethoxypyrimidin-2-yl)thiazolidin-4-one (17)
White solid, yield: 80.7%. Mp: 110-111 °C; IR (KBr): νmax C=O 1742cm-1; 1H NMR (300
MHz, CDCl3); δ 7.70-7.58 (m, 2H, H3,5-PhH), 7.37 (t, 2H, J = 8.0 Hz, H4-PhH, H5-pyrimidin-
2-yl), 5.75 (s, 1H, CH), 4.23 (d, 1H, J = 16.3 Hz, CH2), 3.99 (dd, 1H, J = 14.6 Hz, 2.7 Hz,
CH2), 3.71 (s, 6H, di-OCH3 at C4,6-pyrimidin-2-yl); 13C NMR (75 MHz, CDCl3): δ 171.3
(C=O, pyrimidin-2-yl-C4 & C6), 155.1 (pyrimidin-2-yl-C2), 150.1, 133.6, 133.5, 128.7, 122.8
(Ph-C), 86.9 (pyrimidin-2-yl-C5), 57.6 (OCH3 at C4,6-pyrimidin-2-yl), 54.1 (C2). 29.6 (C5);
MS (ESI): m/z 397.1 (M+H+). HRMS (ESI): (M+H+), calcd. for C15H13ClN4O5S+H+ m/z
397.0373, Found 397.0382.
4.2.18. 3-(5-bromo-4,6-dimethylpyridin-2-yl)-2-(2-chloro-6-nitrophenyl)thiazolidin-4-one
(18)
White solid, yield: 72.6%. Mp: 113-114 °C; IR (KBr): νmax C=O 1698cm-1; 1H NMR (300
MHz, CDCl3); δ 8.03 (s, 1H, H5-PhH), 7.68 (d, 1H, J = 7.9 Hz, H3-pyridin-2-yl), 7.50 (d, 1H,
J = 7.7 Hz, H3-PhH), 7.33 (d, 1H, J = 8.0 Hz, H4-PhH), 7.16 (s, 1H, CH), 4.23 (d, 1H, J = 17.3
Hz, CH2), 3.99 (d, 1H, J = 15.9 Hz, CH2), 2.38 (s, 3H, CH3 at C6-pyridin-2-yl), 2.32 (s, 3H,
CH3 at C4-pyridin-2-yl); 13C NMR (75 MHz, CDCl3): δ 171.2 (C=O), 155.3 (pyridin-2-yl-C6),
149.4, 149.7 (pyridin-2-yl-C4 & Ph-C2), 135.7 (pyridin-2-yl-C2), 132.5, 128.8, 122.4, 119.9
(Ph-C), 114.5 (pyridin-2-yl-C3 & C5), 57.0 (C2). 35.7 (C5) 23.3, 23.7 (CH3 at C4,6-pyridin-2-
yl); MS (ESI): m/z 442.0 (M+). HRMS (ESI): (M+H+), calcd. for C16H13BrClN3O3S +H+ m/z
441.9628, Found 441.9994.
4.2.19. 3-(5-bromo-4,6-dimethylpyridin-2-yl)-2-(2,6-difluorophenyl)thiazolidin-4-one (19)
17

White solid, yield: 80.5%. Mp: 118-119 °C; IR (KBr): νmax C=O 1699 cm-1; 1H NMR (300
MHz, CDCl3); δ 8.03 (s, 1H, H3-pyridin-2-yl), 7.23-7.13 (m, 2H, H3,5-PhH), 6.86 (t, 2H, J =
8.7 Hz, H4-PhH & CH), 4.26 (d, 1H, J = 15.8 Hz, CH2), 3.87 (d, 1H, J = 15.8 Hz, CH2), 2.46
(s, 3H, CH3 at C6-pyridin-2-yl), 2.39 (s, 3H, CH3 at C4-pyridin-2-yl); 13C NMR (75 MHz,
CDCl3): δ 171.0 (C=O), 162.3, 162.2 (Ph-C2 & C6), 159.0, 155.1, 149.2 (pyridin-2-yl-C6, C4
& C2), 129.5 (Ph-C4), 119.6, 118.5 (pyridin-2-yl-C5 & C3), 114.4, 111.8, 111.5 (Ph-C1, C3
& C5) 53.2 (C2), 35.0 (C5), 24.6, 23.7 (CH3 at C4,6-pyridin-2-yl); MS (ESI): m/z 399.1 (M+).
HRMS (ESI): (M+H) +, calcd. for C16H13BrF2N2OS+H+ m/z 398.9978, Found 398.9401.
4.2.20. 3-(5-bromo-4,6-dimethylpyridin-2-yl)-2-(2,6-dimethoxyphenyl)thiazolidin-4-one
(20)
Dark yellow solid, yield: 65.5%. Mp: 115-117 °C; IR (KBr): νmax C=O 1702 cm-1; 1H NMR
(300 MHz, CDCl3); δ 7.90 (s, 1H, H3-pyridin-2-yl), 7.44 (s, 1H, CH), 7.15 (t, 1H, J = 8.3 Hz,
H4-PhH), 6.50 (d, 2H, J = 8.2 Hz, H3,5-PhH), 4.17 (dd, 2H, J = 15.3 Hz, 1.5 Hz, CH2), 3.91-
3.73 (m, 6H, di-OCH3), 2.42 (s, 3H, CH3 at C6-pyridin-2-yl), 2.36 (s, 3H, CH3 at C4-pyridin-2-
yl); 13C NMR (75 MHz, CDCl3): δ 171.3 (C=O), 159.2 (Ph-C2 & C6), 158.9 (pyridin-2-yl-
C6), 151.2 (pyridin-2-yl-C4), 143.6 (pyridin-2-yl-C2), 128.7 (Ph-C4), 116.9 (pyridin-2-yl-C3
& C5), 110.1 (Ph-C1), 105.5 (Ph-C3 & C5), 61.1 (C2), 54.7 (OCH3 at C2,6-Ph), 32.5 (C5),
24.7, 23.8 (CH3 at C4,6-pyridin-2-yl); MS (ESI): m/z 423 (M+). Anal. Calcd for
C18H19BrN2O3S: C, 51.07; H, 4.52; N, 6.62. Found: C, 51.15; H, 4.50; N, 6.59.
4.2.21. 2-(2,6-dimethoxyphenyl)-3-(4,6-dimethoxypyrimidin-2-yl)thiazolidin-4-one (21)
White solid, yield: 72.6%. Mp: 125-126 °C; IR (KBr): νmax C=O 1731cm-1; 1H NMR (300
MHz, CDCl3); δ 7.18-7.12 (m, 2H, H3,5-PhH), 6.53 (d, 2H, J = 8.3 Hz, C4-pyrimidin-2-yl &
H4-PhH ), 5.71 (s, 1H, CH), 4.14 (dd, 2H, J = 15.4 Hz, 0.9 Hz, CH2), 3.86-3.75 (m, 12H,
18

OCH3 at C2,6-PhH & C4,6-pyrimidin-2-yl); 13C NMR (75 MHz, CDCl3): δ 171.4 (C=O), 170.6
(pyrimidin-2-yl-C4 & C6), 158.6 (Ph-C2 & C6), 155.1 (pyrimidin-2-yl-C2), 128.1 (Ph-C4),
108.1 (Ph-C1), 104.5 (Ph-C3 & C5), 78.1 (pyrimidin-2-yl-C5), 58.9 (C2), 56.1 (OCH3 at C2,6-
Ph), 54.8 (OCH3 at C4,6-pyrimidin-2-yl), 31.2 (C5); MS (ESI): m/z 423 (M+). MS (ESI): m/z
378.1 (M+H)+. HRMS (ESI): (M+H)+, calcd. for C17H19N3O5S+H+ m/z 378.1124, Found
378.1052.
4.2.22. 2-(2,6-dimethoxyphenyl)-3-(5-methylisoxazol-3-yl)thiazolidin-4-one (22)
White solid, yield: 86.7%. Mp: 101-102 °C; IR (KBr): νmax C=O 1705 cm-1; 1H NMR (300
MHz, CDCl3); δ 7.21 (t, 1H, J = 8.3 Hz, H4-PhH), 6.97 (s, 1H, H4-isoxazol-3-yl-H), 6.78 (s,
1H, CH), 7.59-6.49 (m, 2H, H3,5-PhH), 4.06 (dd, 2H, J = 15.5 Hz, 1.3 Hz, CH2), 3.92, 3.76 (s,
6H, OCH3 at C2,6-Ph), 2.33 (s, 3H, CH3); 13C NMR (75 MHz, CDCl3): δ 172.2 (C=O), 169.3
(isoxazol-3-yl-C5), 158.1 (isoxazol-3-yl-C3), 157.8, 129.5, 116.5, 104.3 (Ph-C), 95.4
(isoxazol-3-yl-C4), 56.4, 55.8 (OCH3 at C2,6-Ph), 53.7 (C2), 34.4 (C5), 12.5 (CH3 at C5-
isoxazol-3-yl); MS (ESI): m/z 320.9 (M+). HRMS (ESI): (M+H+), calcd. for C15H16N2O4S+H+
m/z 321.0909, Found 321.0919.
4.2.23. 2-(2,6-dichlorophenyl)-3-(9H-purin-6-yl)thiazolidin-4-one (23)
White solid, yield: 65.4 %. Mp: 156-158 °C; IR (KBr): νmax C=O 1630 cm-1; 1H NMR (300
MHz, DMSO); δ 12.72 (br, 1H, NH, purin-6-yl), 8.59 (d, 2H, J = 6.8 Hz, H2,8-purin-6-yl),
7.56 (s, 1H, CH), 7.48-7.45 (m, 1H, H4-PhH), 7.28-7.21 (m, 2H, H3,5-PhH), 4.19 (s, 2H, CH2);
13C NMR (75 MHz, DMSO): δ 171.5 (C=O), 150.8 (purin-6-yl-C2 & C4), 147.3 (purin-6-yl-
C6), 135.1 (purin-6-yl-C8 & Ph-C1), 133.6 (Ph-C2 & C6), 133.0 (Ph-C4), 131.2, 131.0 (Ph-
19

C3 & C5), 129.3 (purin-6-yl-C5), 58.3 (C2), 34.4 (C5); MS (ESI): m/z 367.2 (M+H+). HRMS
(ESI): (M+H+), calcd. for C14H9Cl2N5OS+H+ m/z 365.9983, Found 365.9975.
4.2.24. 2-(2,6-difluorophenyl)-3-(9H-purin-6-yl)thiazolidin-4-one (24)
White solid, yield: 70.2 %. Mp: 147-149 °C; IR (KBr): νmax C=O 1635 cm-1; 1H NMR (300
MHz, DMSO); δ 12.70 (br, 1H, NH, purin-6-yl), 8.61 (d, 2H, J = 10.17 Hz, H2,8-purin-6-yl),
7.39-7.29 (m, 1H, H4-PhH), 7.11 (s, 1H, CH), 7.06 (t, 2H, J = 9.1 Hz, H3,5-PhH), 4.21 (d, 1H,
J = 16.0 Hz, CH2), 4.13 (d, 1H, J = 16.7 Hz, CH2); 13C NMR (75 MHz, DMSO): δ 171.7
(C=O), 161.9 (Ph-C2 & C6), 158.6, 151.0 (purin-6-yl-C2 & C4), 147.6 (purin-6-yl-C6), 131.4
(purin-6-yl-C8), 131.3 (Ph-C4), 116.8 (purin-6-yl-C5), 115.8 (Ph-C1), 112.8, 112.5 (Ph-C3 &
C5), 58.0 (C2), 33.7 (C5); MS (ESI): m/z 334.0 (M+H+). HRMS (ESI): (M+H+), calcd. for
C14H9F2N5OS+H+ m/z 334.0574, Found 334.0581.
4.2.25. 2-(2,6-dichlorophenyl)-3-(4,6-dimethoxypyrimidin-2-yl)thiazolidin-4-one (25)
White solid, yield: 80.8%. Mp: 122-123 °C; IR (KBr): νmax C=O 1741cm-1; 1H NMR (300
MHz, CDCl3); δ 7.31-7.25 (m, 3H, H3,4,5-PhH), 7.16 (t, 1H, J = 8.0 Hz, H5-pyrimidin-2-yl),
5.78 (s, 1H, CH), 4.19 (dd, 1H, J = 15.8 Hz, 1.3 Hz, CH2), 3.92 (d, 1H, J = 15.8 Hz, CH2),
3.79 (s, 6H, di-OCH3 at C4,6-pyrimidin-2-yl); 13C NMR (75 MHz, CDCl3): δ 171.4 (C=O),
170.3 (pyrimidin-2-yl-C4 & C6), 155.3 (pyrimidin-2-yl-C2), 135.7, 134.4, 133.1, 130.7,
129.0, 128.4 (Ph-C), 86.7 (pyrimidin-2-yl-C5), 59.2 (OCH3 at C4,6-pyrimidin-2-yl), 54.1 (C2).
35.3 (C5); MS (ESI): m/z 386.2 (M+). HRMS (ESI): (M+), calcd. for C15H13Cl2N3O3S+ m/z
385.0055, Found 385.0096.
4.2.26. 2-(2-chloro-6-fluorophenyl)-3-(4,6-dimethoxypyrimidin-2-yl)thiazolidin-4-one
(26)
20

White solid, yield: 84.8%. Mp: 124-125 °C; IR (KBr): νmax C=O 1737cm-1; 1H NMR (300
MHz, CDCl3); δ 7.29-7.19 (m, 2H, H3,5-PhH), 7.01-6.94 (m, 2H, H5-pyrimidin-2-yl & H4-
PhH), 5.78 (s, 1H, CH), 4.18 (d, 1H, J = 15.9 Hz, CH2), 3.91 (d, 1H, J = 15.8 Hz, CH2), 3.80
(s, 6H, di-OCH3 at C4,6-pyrimidin-2-yl); 13C NMR (75 MHz, CDCl3): δ 171.4 (C=O), 170.1
(pyrimidin-2-yl-C4 & C6), 162.3 (Ph-C2), 159.0 (pyrimidin-2-yl-C2), 155.3, 133.0, 129.2,
125.5 (Ph-C), 86.5 (pyrimidin-2-yl-C5), 58.0 (OCH3 at C4,6-pyrimidin-2-yl), 54.1 (C2). 30.9
(C5); MS (ESI): m/z 370.2 (M+H+). HRMS (ESI): (M+H+), calcd. for C15H13ClFN3O3S+H+
m/z 370.0428, Found 370.0423.
4.2.27. 2-(2,6-difluorophenyl)-3-(4,6-dimethoxypyrimidin-2-yl)thiazolidin-4-one (27)
White solid, yield: 90.2%. Mp: 117-118 °C; IR (KBr): νmax C=O 1737cm-1; 1H NMR (300
MHz, CDCl3); δ 7.26-7.19 (m, 1H, H5-pyrimidin-2-yl), 6.91 (t, 3H, J = 8.6 Hz, H3,4,5-PhH),
5.78 (s, 1H, CH), 4.23 (d, 1H, J = 15.2 Hz, CH2), 4.16 (d, 1H, J = 14.2 Hz, CH2), 3.84 (s, 6H,
di-OCH3 at C4,6-pyrimidin-2-yl); 13C NMR (75 MHz, CDCl3): δ 171.4 (C=O), 169.9
(pyrimidin-2-yl-C4 & C6), 162.0, 161.9 (Ph-C2 & C6), 158.6 (pyrimidin-2-yl-C2), 129.4,
112.0, 111.9, 111.6 (Ph-C), 86.5 (pyrimidin-2-yl-C5), 54.0 (OCH3 at C4,6-pyrimidin-2-yl),
53.2 (C2). 34.6 (C5); MS (ESI): m/z 354.2 (M+H+). HRMS (ESI): (M+H+), calcd. for
C15H13F2N3O3S+H+ m/z 354.0724, Found 354.0718.
4.3. General procedure for the preparation of compounds (28-33) from pyridyl-2-yl/4-
cyanophenyl hydrazine
An equimolar mixture of 2,6-dihalo-substituted benzaldehyde (1.0 mmol) and aryl/heteroaryl
hydrazine (1.0 mmol) was refluxed in toluene using a Dean–Stark trap for 5 hours. The
solvent was evaporated under reduced pressure by using rotary evaporator. The solid
hydrazone intermediates were added to the excess mercaptoacetic acid (5 mmol), and then
21

heated at 60 °C for 12 hours until reaction was completed, as shown by TLC. Ethyl acetate (5
ml) was added, the organic layer was washed with saturated sodium bicarbonate (3 X 20 ml)
and water (1 X 10 ml), dried with sodium sulphate, and concentrated to give an oil. The oil
was purified by column chromatography on silica gel using hexane/ethyl acetate as eluent.
4.3.1. 2-(2,6-dichlorophenyl)-3-(pyridin-2-ylamino)thiazolidin-4-one (28)
White solid, yield: 75.9%. Mp: 118-119 °C; IR (KBr): νmax -NH- 3350, C=O 1699 cm-1; 1H
NMR (300 MHz, CDCl3); δ 8.21 (d, 1H, J = 4.4 Hz, H6-pyridin-2-yl), 7.59-7.54 (m, 1H, H4-
pyridin-2-yl ), 7.40 (d, 1H, J = 7.7 Hz, H4-PhH), 7.32-7.20 (m, 2H, H3,5-PhH), 7.03 (s, 1H,
CH), 6.87 (t, 1H, J = 5.0 Hz, H5-pyridin-2-yl), 6.67 (d, 1H, J = 8.2 Hz, H3-pyridin-2-yl), 6.43
(br, 1H, NH), 3.85 (s, 2H, CH2); 13C NMR (75 MHz, CDCl3): δ 169.9 (C=O), 157.1, 148.3,
138.1, 136.4, 134.4, 132.1, 130.7, 130.2, 128.9, 117.1, 107.4 (aromatic-C), 57.9 (C2), 30.8
(C5); MS (ESI): m/z 340.4 (M+). HRMS (ESI): (M+H+), calcd. for C14H11Cl2N3OS+H+ m/z
340.0078, Found 340.0081.
4.3.2. 2-(2-chloro-6-fluorophenyl)-3-(pyridin-2-ylamino)thiazolidin-4-one (29)
White solid, yield: 95.8%. Mp: 135-136 °C; IR (KBr): νmax -NH- 3401, C=O 1707 cm-1; 1H
NMR (300 MHz, CDCl3); δ 8.21 (d, 1H, J = 4.4 Hz, H6-pyridin-2-yl), 7.59 (t, 1H, J = 7.3 Hz,
H4-pyridin-2-yl ), 7.20 (d, 1H, J = 7.9 Hz, H5-PhH), 7.10 (t, 1H, J = 10.6 Hz, H3-PhH), 6.87 (t,
1H, J = 5.3 Hz, H4-PhH), 6.73-6.63 (m, 3H, H3,5-pyridin-2-yl & CH-thiazolidin-4-one), 6.50
(br, 1H, NH), 3.91 (d, 1H, J = 15.6 Hz, CH2), 3.77 (d, 1H, J = 15.6 Hz, CH2); 13C NMR (75
MHz, CDCl3): δ 169.9 (C=O), 163.9, 157.3, 148.3, 138.2, 134.7, 130.6, 127.5, 125.9, 124.8,
115.8, 107.2 (aromatic-C), 56.7 (C2), 29.7 (C5); MS (ESI): m/z 324.0 (M+H+). HRMS (ESI):
(M+H+), calcd. for C14H11ClFN3OS+H+ m/z 324.0374, Found 324.0906.
22

4.3.3. 2-(2,6-difluorophenyl)-3-(pyridin-2-ylamino)thiazolidin-4-one (30)
White solid, yield: 93.6%. Mp: 128-129 °C; IR (KBr): νmax -NH- 3345, C=O 1699 cm-1; 1H
NMR (300 MHz, CDCl3); δ 8.20 (d, 1H, J = 4.6 Hz, H6-pyridin-2-yl), 7.59 (t, 1H, J = 7.4 Hz,
H4-pyridin-2-yl ), 7.36-7.31 (m, 1H, H4-PhH), 6.96-6.83 (m, 3H, H3,5-PhH & H5-pyridin-2-yl),
6.68 (d, 1H, J = 8.2 Hz, H3-pyridin-2-yl), 6.58 (br, 1H, NH), 7.41 (s, 1H, CH), 3.94 (d, 1H, J =
15.6 Hz, CH2), 3.77 (d, 1H, J = 15.5 Hz, CH2); 13C NMR (75 MHz, CDCl3): δ 169.8 (C=O),
163.6, 163.5, 158.7, 157.3, 148.2, 138.1, 117.0, 112.3, 111.2, 111.8, 107.1 (aromatic-C), 52.5
(C2), 29.8 (C5); MS (ESI): m/z 308.2 (M+H+). HRMS (ESI): (M+H+), calcd. for
C14H11F2N3OS+H+ m/z 308.0669, Found 308.0616.
4.3.4. 4-(2-(2,6-dichlorophenyl)-4-oxothiazolidin-3-ylamino)benzonitrile (31)
White solid, yield: 55.2 %. Mp: °C; IR (KBr): νmax -NH- 3421, C=O 1699, CN 2220 cm-1; 1H
NMR (300 MHz, CDCl3); δ 7.57 (d, 2H, J = 8.1 Hz, H2,6-Ph-4-yl), 7.43-7.24 (m, 2H, H3,5-Ph-
1-yl ), 6.84 (d, 3H, J = 8.2 Hz, H4-Ph-1-yl & H3,5-Ph-4-yl), 6.09 (s, 1H, CH), 3.91 (d, 1H, J =
16.0 Hz, CH2), 3.85 (d, 1H, J = 15.8 Hz, CH2); 13C NMR (75 MHz, CDCl3): δ 169.2 (C=O),
148.1, 135.3, 132.8, 132.7, 130.6, 129.7, 129.4, 128.1, (aromatic-C), 118.1 (CN), 112.2, 103.2
(aromatic-C), 56.3 (C2), 29.6 (C5); MS (ESI): m/z 364.0 (M+). HRMS (ESI): (M+H+), calcd.
for C16H11Cl2N3OS+H+ m/z 364.0078, Found 364.0049.
4.3.5. 4-(2-(2-chloro-6-fluorophenyl)-4-oxothiazolidin-3-ylamino)benzonitrile (32)
White solid, yield: 75.8 %. Mp: °C; IR (KBr): νmax -NH- 3421, C=O 1699, CN 2223 cm-1; 1H
NMR (300 MHz, CDCl3); δ 7.51 (d, 2H, J = 8.5 Hz, H2,6-Ph-4-yl), 7.34-7.20 (m, 2H, H3,5-Ph-
1-yl ), 7.10, (t, 1H, J = 10.5 Hz, H4-Ph-1-yl), 6.79 (d, 2H, J = 8.5 Hz, H3,5-Ph-4-yl), 6.62 (s,
1H, NH), 6.42 (s, 1H, CH), 3.91 (d, 1H, J = 15.7 Hz, CH2), 3.74 (d, 1H, J = 15.8 Hz, CH2);
13C NMR (75 MHz, CDCl3): δ 170.4 (C=O), 159.5, 149.3, 133.8, 130.9, 130.7, 126.1, 124.3,
23

124.1, (aromatic-C), 119.2 (CN), 115.7, 115.4, 113.1, 104.1 (aromatic-C), 56.3 (C2), 29.6
(C5); MS (ESI): m/z 347.0 (M+). HRMS (ESI): (M+H+), calcd. for C16H11ClFN3OS +H+ m/z
348.0374, Found 348.0381.
4.3.6. 4-(2-(2,6-difluorophenyl)-4-oxothiazolidin-3-ylamino)benzonitrile (33)
White solid, yield: 91.2 %. Mp: °C; IR (KBr): νmax -NH- 3398, C=O 1705, CN 2221 cm-1; 1H
NMR (300 MHz, CDCl3); δ 7.56 (d, 2H, J = 8.4 Hz, H2,6-Ph-4-yl), 7.39-7.31 (m, 1H, H3-Ph-
1-yl ), 6.99 (t, 2H, J = 8.9 Hz, H4,5-Ph-1-yl ), 6.83 (d, 2H, J = 8.4 Hz, H3,5-Ph-4-yl), 6.31 (s,
1H, NH), 6.21 (s, 1H, CH), 3.95 (d, 1H, J = 15.3 Hz, CH2), 3.75 (d, 1H, J = 15.7 Hz, CH2);
13C NMR (75 MHz, CDCl3): δ 170.2 (C=O), 162.6, 162.5, 159.3, 149.3, 133.7, 131.1, 131.0
(aromatic-C), 119.1 (CN), 115.2, 114.8, 113.1, 112.4, 104.2 (aromatic-C), 52.2 (C2), 29.6
(C5); MS (ESI): m/z 331.0 (M+). HRMS (ESI): (M+H+), calcd. for C16H11F2N3OS+H+ m/z
332.0669, Found 332.0667.
4.4. In vitro HIV-RT kit assay
The enzymatic HIV-RT inhibition assay was performed by using an RT assay kit following
manufacturer’s instructions (Roche Diagnostics GmbH, Germany).10,18 Briefly, the reaction
mixture containing template/primer complex, 2'-deoxy-nucleotide-5'-triphosphates (dNTPs)
and reverse transcriptase (RT) enzyme in the lysis buffer was incubated for 1 hour at 37 °C
and subsequently, the reaction mixture was transferred to streptavidin-coated microtitre plate
(MTP). The biotin labeled dNTPs that are incorporated in the template due to the activity of
RT bind to streptavidin. The unbound dNTPs were washed using wash buffer and anti-
digoxigenin-peroxidase (anti-DIG-POD) was added onto MTP. The DIG-labeled dNTPs
incorporated in the template was bound to anti-DIG-POD antibody. The unbound anti-DIG-
POD was washed and the peroxide substrate (ABST) was added to the MTP. A colored
24

reaction product was produced during the cleavage of the substrate catalyses by a peroxide
enzyme. The absorbance of the sample was determined at OD405 nm using microtiter plate
ELISA reader. The resulting color intensity is directly proportional to the actual RT activity.
The percentage inhibitory activity of RT inhibitors was calculated by comparing with a
sample that does not contain an inhibitor. The percentage inhibition was calculated by the
formula as given;
4.5. In vitro Anti-HIV assay
All the thiazolidin-4-one compounds discussed here were evaluated for in vitro antiviral
activity against HIV-1IIIB, HIV-1ADA5, HIV-1UG070 and HIV-1VB59 in TZM-bl cells as per
previously reported procedure.10,19
Cell line and viral Stocks
Cell Line: TZM-bl (JC53BL-13) cells were obtained from the NIH AIDS Research and
Reference Reagent Programme (ARRRP), NIH, USA. These are genetically modified HeLa
cells, which express CD4, CXCR4, and CCR5 receptors. In addition, the cells also contain
Tat-responsive reporter genes encoding firefly luciferase (Luc) and Escherichia coli β-
galactosidase under the regulatory control of an HIV-1 long terminal repeat. The cells were
maintained in Dulbecco’s modified Eagle’s medium DMEM (Gibco, USA) containing 10%
heat inactivated fetal bovine serum (Morgate, Australia), HEPES (Gibco, USA), penicillin and
25

streptomycin (Gibco, USA) and gentamicin (Sigma, USA). Cultures were incubated at 37 oC
in a humidified 5% CO2 atmosphere and used when 80% confluency was achieved.
Viral Stocks: Lab adapted HIV-1IIIB (X4, subtype B), HIV-1ADA5 (R5, subtype B) and the
primary isolates HIV-1UG070 (X4, subtype D) were obtained from NIH ARRRP, whereas R5
tropic isolate HIV-1VB59 (subtype C) is an indian isolate from the National AIDS Research
Institute, Pune. The viruses were grown in PHA-P (5µg/ml) (Sigma Aldrich, USA) activated
peripheral blood mononuclear cells (PBMC) derived from healthy donors. Virus production
was quantified in cell culture supernatants by HIV p24 antigen detection kit (Advanced
Bioscience Laboratories Inc. Kit, USA). Aliquots of cell-free viral culture supernatants were
centrifuged, filtered and stored at -70 oC. The TCID50 (50% Tissue culture infective dose) of
each virus stock was determined in the TZM-bl cells using Spearman Karber formula.
4.5.1. Cytotoxicity assay in TZM-bl cell line
The TZM-bl cells (1×104 cells/well) were seeded in a 96-well plate and incubated overnight at
37 oC with 5% CO2 atmosphere. Next day, the medium was replaced with 100 µl of two fold
dilution series of each compound in quadruplicate and the plate was incubated further. After
48 hours, cell viability was determined by measuring the mitochondrial-dependent conversion
of the MTT salt (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) (Sigma
Aldrich, USA) to a coloured formazan product and the CC50 (concentration showing 50%
toxicity) was calculated. In this assay, the measured absorbance is proportional to the viable
cell number and inversely to the degree of cytotoxicity. For each compound the results are
represented as mean of three independent assays.
26

4.5.2. Cell associated Anti-HIV-1 assay
The TZM-bl cells (1×104/well) were seeded in a 96-well plate and incubated overnight at 37
oC with 5% CO2 atmosphere. On the subsequent day, the cells were infected with 400 TCID50
HIV-1 stock and incubated for 2 hours at 37 oC with 5% CO2 atmosphere. Serial dilutions of
each compound showing more than 50% viability were then added in duplicates onto the cells.
Anti-HIV-1 activity was determined after 48 hours of incubation by measuring the relative
luminescence units (RLU) using the Britelite Plus substrate (Perkin Elmer). In this assay, RLU
is directly proportional to the number of virus particles and inversely to the percent of
inhibition. The concentration showing 50% inhibition (IC50) was determined using Luc
software. The results were interpreted as mean of two independent assays. Nevirapine was
used as a positive control.
4.6. Docking studies
The Autodock 4.0 program was used to locate the appropriate binding orientations and
conformations of thiazolidin-4-ones of HIV-1 RT inhibitors.16 Autodock is an extensively
used automated procedure for predicting the interactions of small molecules, such as peptides,
enzyme inhibitors, and drugs, to macromolecules, such as proteins, enzymes, antibodies,
DNA, and RNA. The structure of the non-nucleoside binding site (NNBS) of RT was taken
from the crystal structure of HIV-1 RT in complex with its inhibitor TMC278 filed in the
Brookhaven Protein Data Bank with the entry code 2ZD1.17 The molecular structures of
thiazoldin-4-one derivatives were built using the SKETCH option in Sybyl 7.3 version.
Geometry optimizations of all chemical structures were carried out using the Tripos force field
with a distance-dependent dielectric and the Powell conjugate gradient algorithm. Gasteiger-
Hückel charges were used. Prior to docking studies, all water molecules, TMC278 (ligand)
27

and magnesium ion were removed from the original Protein Data Bank files. Polar hydrogen
atoms were added and gasteiger charges, atomic solvation parameters and fragmental volumes
were allocated to the protein. All torsions in thiazolidin-4-ones were treated as flexible by
allowing them to rotate freely. The grid map calculation was performed using Autogrid 4.0
program, with a dimension of 60 X 60 X 60 points, a grid-point spacing of 0.375 Å and the
maps were centered on the ligand binding site. The Lamarckian genetic algorithm (LGA) in
Autodock 4.0 was used to explore the energy landscape. The hybrid search technique consists
of a global optimizer modified from a genetic algorithm with 2-point crossover, random
mutation, and a local optimizer with a Solis and Wets algorithm. A docking box with a
dimension of 60 X 60 X 60 points with a grid spacing of 0.375 Å was defined in the
calculations. Random conditions were used in the settings of seed, initial quaternion,
coordinates, and torsions. A 0.2 Å step was used for translation and a 25-degree was used for
quaternion and torsion. The maximum number of energy evaluation with 250 000 and the
maximum number of generations with 27 000 was applied. The rate of gene mutation and
crossover was set to 0.02 and 0.8, respectively. The number of cycles was set to 100. As a
result, a total of 100 docking configurations were obtained in each docking calculation. A
“preferable” docking configuration was chosen based on the lowest empirical binding free
energy and the most frequent cluster.
Acknowledgements
This work is supported by ICMR project GAP 0044. We thank CSIR, New Delhi for
supporting Dr. V. Murugesan for Senior Research fellowship. The authors thank the Director,
CDRI for support and the SAIF for the spectral data. CDRI manuscript number: 18/2014/SBK
28

References and notes
1. De Béthune, M. P. Antiviral Res. 2010, 85, 75.
2. Ammaranond, P.; Sanguansittianan, S. Fundam Clin Pharmacol. 2012, 26, 146.
3. Adams, J.; Patel, N.; Mankaryous, N.; Tadros, M.; Miller, C. D. Ann Pharmacother. 2010,
44, 157.
4. Hawkins, T. Antiviral Res. 2010, 85, 201.
5. Menéndez-Arias, L. Antiviral Res. 2010, 85, 210.
6. Deeks, E. D.; Keating, G. M. Drugs. 2008, 68, 2357.
7. Das, K.; Bauman, J. D.; Clark, A. D. Jr.; Frenkel, Y. V.; Lewi, P. J.; Shatkin, A. J.; Hughes,
S. H.; Arnold, E. Proc Natl Acad Sci U S A. 2008, 105, 1466.
8. Barreca, M. L.; Balzarini, J.; Chimirri, A.; Clercq, E. D.; Luca, L. D.; Holtje, H. D.; Holtje,
M.; Monforte, A. M.; Monforte, P.; Pannecouque, C.; Rao, A.; Zappala, M. J Med Chem.
2002, 45, 5410.
9. Rao, A.; Balzarini, J.; Carbone, A.; Chimirri, A.; Clercq, E. D.; Monforte, A. M.; Monforte,
P.; Pannecouque, C.; Zappalà, M. Antivir Res. 2004, 63, 79.
10. Murugesan, V.; Tiwari, V. S.; Saxena, R.; Tripathi, R.; Paranjape, R.; Kulkarni, S.;
Makwana, N.; Suryawanshi, R.; Katti, S. B. Bioorg Med Chem. 2011, 19, 6919.
11. Janssen, P. A.; Lewi, P. J.; Arnold, E.; Daeyaert, F.; De Jonge, M.; Heeres, J.; Koymans,
L.; Vinkers, M.; Guillemont, J.; Pasquier, E.; Kukla, M.; Ludovici, D.; Andries, K.; De
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