Design, synthesis, and structure-activity relationship of a novel series of 2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazolidinylidenemethyl)furans as HIV-1 entry inhibitors

Article abstract of DOI:10.1021/jm900450n

We previously identified two small molecules targeting the HIV-1 gp41, N-(4-carboxy-3-hydroxy)phenyl-2,5-dimethylpyrrole 12 (NB-2) and N-(3-carboxy-4-chloro)phenylpyrrole 13 (NB-64), that inhibit HIV-1 infection at low micromolar levels. On the basis of m

Full text of DOI:10.1021/jm900450n

J. Med. Chem. 2009, 52, 7631–7639 7631
DOI: 10.1021/jm900450n
Design, Synthesis, and Structure-Activity Relationship of a Novel Series of 2-Aryl
5-(4-Oxo-3-phenethyl-2-thioxothiazolidinylidenemethyl)furans as HIV-1 Entry Inhibitors^
Alan R. Katritzky,*,§ Srinivasa R. Tala,§ Hong Lu,‡ Anatoliy V. Vakulenko,§ Qi-Yin Chen,§ Jothilingam Sivapackiam,§
Keyur Pandya,§ Shibo Jiang,*,‡ and Asim K. Debnath*,‡
§Center for Heterocyclic Compounds, Department of Chemistry, University of Florida, Gainesville, Florida 32611, and
‡Lindsley F. Kimball Research Institute, New York Blood Center, New York 10065
Received April 7, 2009
We previously identified two small molecules targeting the HIV-1 gp41, N-(4-carboxy-3-hydroxy)phenyl-
2,5-dimethylpyrrole 12 (NB-2) and N-(3-carboxy-4-chloro)phenylpyrrole 13 (NB-64), that inhibit HIV-1
infection at low micromolar levels. On the basis of molecular docking analysis, we designed a series of
2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazolidinylidenemethyl)furans. Compared with 12 and 13, these
compounds have bigger molecular size (437-515 Da) and could occupy more space in the deep hydro-
phobic pocket on the gp41 NHR trimer. Fifteen 2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazolidinylidene-
methyl)furans (11a-o) were synthesized by Suzuki-Miyaura cross-coupling followed by a Knoevenagel
condensation and tested for their anti-HIV-1 activity and cytotoxicity on MT-2 cells. We found that all
15 compounds had improved anti-HIV-1 activity and 3 of them (11a, 11b, and 11d) exhibited inhibitory
activity against replication of HIV-1IIIB and 94UG103 at <100 nM range, more than 20-fold more potent
than 12 and 13, suggesting that these compounds can serve as leads for development of novel small molecule
HIV fusion inhibitors.
Introduction
gp120 to the CD4 molecule and a coreceptor, CCR5 or
CXCR4, causing a series of conformational changes of the
Env transmembrane subunit gp41.7 The fusion peptide lo-
cated at the N-terminus of gp41 inserts into the target cell
membrane, followed by association between the N- and
C-terminal heptad repeats (NHR and CHR, respectively) to
form a six-helix bundle core structure, bringing the viral
envelope and target cell membrane into proximity.8,9 Peptides
derived from the gp41 CHR region can interact with the viral
gp41 NHR region and block fusogenic core formation, resulting
in potent inhibition of HIV-1 fusion with the target cells.10-14
One of the CHR-peptides, enfuvirtide (T-20), was licensed by
the U.S. FDA as the first member of a new class of anti-HIV
drugs, HIV fusion inhibitors for treating HIV/AIDS patients
whohavefailedtorespondtoRTIandPI.15,16 However, clinical
application of the enfuvirtide is limited because of its lack of oral
availability, induction of drug resistance, and high cost of
production.9 Therefore, it is critical to develop orally available
non-peptide small-molecule fusion inhibitors.
So far, over 60 million people have been infected with HIV
and of these more than 25 million have died of AIDS.1,2 As of
March of 2009, the U.S. Food and Drug Administration (FDA)
has licensed 28 anti-HIV drugs, including 15 reverse transcrip-
tase inhibitors (RTIa) and 10 protease inhibitors (PI) (http://
www.hivandhepatitis.com/hiv_and_aids/hiv_treat.html). Appli-
cation of these antiretroviral drugs in combination, designated as
highly active antiretroviral therapy (HAART), has led to signifi-
cant reduction of mobility and mortality of HIV/AIDS.3 How-
ever, an increasing number of HIV/AIDS patients fail to respond
to current antiretroviral therapeutics because of the emergence of
multidrug resistance to RTIs and PIs and serious adverse side
effects.4-6 Therefore, there is an urgent need to develop novel anti-
HIV therapeutics targeting different steps of HIV replication
cycle, particularly the HIV fusion and entry events.
HIV type 1 (HIV-1) entry into the host cell is initiated by
binding of its envelope glycoprotein (Env) surface subunit
Using a fluorescence-linked immunoabsorbance assay with
a conformation-specific monoclonal antibody (mAb) NC-1,17
we previously identified two small molecules, N-(4-carboxy-
3-hydroxy)phenyl-2,5-dimethylpyrrole 12 (NB-2) and N-(3-
carboxy-4-chloro)phenylpyrrole 13 (NB-64), which inhibit
HIV-1 fusion and gp41 six-helix bundle formation at low
micromolar levels (Figure 1).18 Molecular docking analysis
of these two compounds indicates that 12 and 13 each
occupy only a part of the deep hydrophobic pocket on the
gp41 NHR trimer.18 We therefore reasoned that new lead
compounds with improved anti-HIV-1 potency could be de-
signed by increasing the molecular size so that they would
occupy more space in the hydrophobic pocket. We now
report the design and synthesis of 15 derivatives of 2-aryl
^ Dedicated to Professor Corwin Hansch on his 91st anniversary in
appreciation of his pioneering achievements.
*To whom correspondence should be addressed. For A.R.K.: phone,
352-392-0554; fax, 352-392-9199; e-mail:, Katritzky@chem.ufl.edu. For
S.J. and A.K.D.: phone, (212) 570-3373; fax, (212) 570-3168; e-mail,
sjiang@nybloodcenter.org (SJ) and adebnath@nybloodcenter.org
(AKD).
a Abbreviations: CHR, C-terminal heptad repeat; Env, envelope glycopro-
tein; NHR, N-terminal heptad repeat; XTT, sodium 30-[1-(phenylamino)-
carbonyl]-3,4-tetrazolium-bis(4-methoxy-6-nitro)bezenesulfonic acid hydrate;
RTI, reverse transcriptase inhibitors; PI, protease inhibitors; HAART, highly
active antiretroviral therapy; mAb, monoclonal antibody; ELISA, enzyme-
linked immunosorbent assay; THF, tetrahydrofuran; LDA, lithium diisopro-
pylamide; i-PrMgCl, isopropylmagnesium chloride; EtOH-DME, ethyl
alcohol-ethylene glycol dimethyl ether; RCSB, Research Collaboratory for
Structural Bioinformatics; MOI, multiplicity of infection; PMS, phenazine
methosulphate.
r
2009 American Chemical Society
Published on Web 09/11/2009
pubs.acs.org/jmc

7632 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23
Katritzky et al.
5-(4-oxo-3-phenethyl-2-thioxothiazolidinylidenemethyl)furan
(11a-o) with molecular weight ranging from 437 to 515 Da.
We evaluated their antiviral activity against HIV-1IIIB and their
cytotoxicity on MT-2 cells that were used in the viral infection
assay. Strikingly, all 15 compounds exhibited improved anti-
HIV-1 activity and 3 of them (11a, 11b, and 11d) inhibited
replication by laboratory-adapted and primary HIV-1 strains
with EC50 (effective concentration for 50% inhibition) ranging
from 44 to 99 nM and SI (selectivity index) in the range of
330-440. These results suggest that 2-aryl 5-(4-oxo-3-phen-
ethyl-2-thioxothiazolidinylidenemethyl)furans show great po-
tential for further development as novel small molecule anti-
HIV therapeutics targeting gp41.
Figure S1 of Supporting Information). The best compound is
11l, which has the lowest EC50 (42 nM) and highest SI (915),
and the worst compound is 11f, with an EC50 of 1.28 μM and a
SI of 12. Although compound 11i had an appreciable potency
(EC50 = 0.32 μM), it had high cytotoxicity (CC50 = 3.29 μM),
resulting in a SI of 10.
Fourpossibleisomers need tobe considered for compounds
11a-o: A, B, C, and D (Figure 2). The interconvertions ATB
and CTD should be easy as they proceed by rotation about
single bonds. The interconversions of ATC and BTD will be
more difficult because they need rotation about double bonds.
We also expect that isomers A and B will be more stable than
C and D because of the cross space interactions. To examine
the possible intervention of the unstable product isomers C
and D, we tested the anti-HIV-1IIIB activity of these com-
pounds under exclusion of light. As shown in Table 3, there
was no substantial difference in antiviral activity of the
compounds tested under dark and light conditions. This
excludes the possibility that light-activation to give isomers
C and D is the origin for their bioactivity.
We also tested the inhibitory activity of these compounds
on the replication of a representative primary HIV-1 isolate
94UG103 (R5X4, clade A) in PBMCs. All 15 compounds
effectively inhibited HIV-1 94UG103 replication similarly in a
dose-dependent manner with the EC50 ranging from 0.07 to
4 μM (Table 3 and Figure S1). The best compounds are 11a,
11b, and 11d, which have the lowest EC50 (70-74 nM). This
result suggests that these compounds are much more potent
than 12 and 13 in inhibiting infection by primary HIV-1.18
Molecular docking analysis revealed that the phenethyl group
of compound 11d filled the space in the deep hydrophobic
pocket of gp41 formed by the NHR trimer (Figure 3), pre-
viously observed to be unoccupied by 13.
Results and Discussion
A total of 15 2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazoli-
dinylidenemethyl)furans (11a-o) were synthesized by Suzuki-
Miyaura cross-coupling, followed by a Knoevenagel condensa-
tion. Their molecular weights ranged from 436 to 515 Da.
We tested the inhibitory activity of these compounds on the
replication of the laboratory-adapted HIV-1 strain IIIB, a
prototype of the X4 strain, in MT-2 cells that express CD4
and the coreceptor CXCR4 by measuring p24 production in an
enzyme-linked immunosorbent assay (ELISA) as previously
described.18 All 15 compounds significantly inhibited HIV-1
replication in a dose-dependent manner with the EC50 (effective
concentration for 50% inhibition) ranging from 0.042 to
1.3 μM. We also tested the cytotoxicity of these compounds
on MT-2 cells using an XTT {sodium 30-[1-(phenylamino)-
carbonyl]-3,4-tetrazolium-bis(4-methoxy-6-nitro)bezenesulfo-
nic acid hydrate} assay.19 The CC50 (concentration causing
50% cytotoxicity) values ranged from 3.29 to 78.29 μM, and
their selectivity indexes (SI) ranged from 10 to 915 (Table 3 and
This may explain why 11d has ∼25-fold greater potency
than13. In thecurrentserieswekept the phenethylgroupfixed
and substituted the phenyl ring attached to the furan ring to
derive
a comprehensive structure-activity relationship
(SAR). We showed previously that the carboxylic acid group
in this phenyl ring is essential for anti-HIV-1 activity, and
hence, it was retained throughout. It appears that the com-
pound 11a with no substituent at the 2 and 6 positions of the
phenyl ring showed highly potent activity with reasonably
good selectivity index. Addition of hydrophobic substituent at
R (11b-d) had moderate effects and reduced the anti-HIV-1
activity by about 2- to 4-fold. However, bromine substitution
Figure 1. Chemical structures of 12 and 13.
Figure 2. Possible E-Z isomers of compounds 11a-o.

Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23 7633
Figure 3. Docking of 11d in the gp41 hydrophobic cavity. (A) The stereoview of 11d docked in the hydrophobic cavity showing possible
interactions with the neighboring hydrophobic and charged residue K574. (B) Surface representation of the gp41 core with bound ligand 11d.
The compound docked inside the cavity. The negatively charged COOH group is pointing toward the positively charged area contributed by
K574.
Scheme 1. Synthesis of 2-Ethyl-5-bromobenzoic Acid 2ca
Scheme 2. Synthesis of 3-Bromo-2-chlorobenzoic Acid 2ja
a Reagents and conditions: (a) Br2/iron (powder), room temperature
a Reagents and conditions: (a) KMnO4, H2O, reflux, 24 h, 49%.
to 50 ꢀC, 14%.
HIV-1 fusion inhibitors with improved anti-HIV-1 potency and
reduced cytotoxicity for further preclinical evaluation.
(11i) hada moresevere effect probably due tostericeffectsand
the activity dropped by ∼7-fold. The most marked adverse
effect on activity was demonstrated by electron donating
substituents OH and OCH3, where the anti-HIV-1 activity
dropped by ∼30-fold. Introduction of a nitrogen atom in the
ring in the unsubstituted 11a yielded the most active inhibitor
(11l EC50= 42 nm, but when chlorine or bromine was
substituted at the R position (11m and 11n) or a methyl group
was introduced at the R0 position (11o), a substantial decrease
in activity was observed. Interestingly moving the methyl
group of 11b to the R0 position yielded 11h, which was more
active than 11b. However, similar moves for chlorine and
fluorine hadmarginal effects. This SAR analysis willhelpus in
further modifying the most active analogues obtained from
the current study.
Chemistry
Synthesis of Aryl(heteryl) Bromide/Iodides 2c, 2j, 2n, 2o.
2-Ethyl-5-bromobenzoic acid 2c was isolated in 14% yield by
treatment of 2-ethylbenzoic acid 1c with bromine in the
presence of iron powder at room temperature for 48 h and
at 55 ꢀC for 2 h (Scheme 1).
3-Bromo-2-chlorobenzoic acid 2j33 was synthesized in
49% yield by the oxidation of 3-bromo-2-chlorotoluene
with KMnO4 in water for 24 h at reflux temperature
(Scheme 2).
2-Bromo-5-iodonicotinic acid 2n was synthesized by treat-
ment of 2,5-dibromo-3-picoline 1n with isopropylmagne-
sium chloride in the presence of I2 in tetrahydrofuran
(THF) at room temperature for 24 h, followed by oxidation34
using KMnO4 in water for 48 h at reflux temperature
(Scheme 3).
4-Methyl-5-iodonicotinic acid 2o was synthesized from 3-
bromo-5-(4,4-dimethyl-1,3-oxazolin-2-yl)pyridine 1o. First,
3-bromo-5-(4,4-dimethyl-1,3-oxazolin-2-yl)pyridine 1o20 was
In the next stage, we will further evaluate the inhibitory
activities of these 15 compounds on the HIV-1-mediated
cell-cell fusion and cytopathic effect as well as the gp41 six-
helix bundle formation. These data in combination with anti-
HIV-1 activity and cytotoxicity well refine the structure-activity
relationship, which will be used to design new, small molecule

7634 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23
Katritzky et al.
treated with MeI in the presence of lithium diisopropylamide
(LDA) at -78 ꢀC for 1 h and for 1 h at room temperature to
afford 3-bromo-4-methyl-5-(4,4-dimethyl-1,3-oxazolin-2-yl)-
pyridine 3o in 76% yield. Reaction of 3-bromo-4-methyl-
5-(4,4-dimethyl-1,3-oxazolin-2-yl)pyridine 3o with isopropyl-
magnesium chloride in the presence of I2 in THF at room
temperature for 24 h followed by hydrolysis gave 4-methyl-
5-iodonicotinic acid 2o (Scheme 4).
was treated with CS2 in diethyl ether at 0-5 ꢀC followed by
treatment with chloroacetic acid under reflux for 1 h to give
3-phenethyl-2-thioxothiazolidin-4-one
(Scheme 5).
6 in 56% yield
Synthesis of 2-Aryl 5-Formylfurans 8. 2-Aryl 5-formylfur-
ans 8 were synthesized by palladium-catalyzed Suzu-
ki-Miyaura cross-coupling22,23 of aryl(heteryl) bromide/
iodide with aryl(heteryl) boronic acids. 2-Aryl 5-formylfur-
ans 8a-o were obtained (i) by coupling of 5-formyl-2-
furylboronic acid 7 with corresponding aryl(heteryl) bro-
mides/iodides 2b-d,f-o to give 8b-d,f-o (28-80% yield)
by a modified literature method24 or (ii) by reaction of
5-bromofuran-2-carboxaldehyde 9 with 3-carboxyphenyl-
boronic acid 10a,e by a slight modifications to Bumagin’s
method25 to give 8a,e (59-89% yield) (Scheme 6, Table 1).
Suzuki-Miyaura coupling did not occur when the bromo
analogues of 2i, 2n, 2o were used instead of aryl(heteryl)
iodides 2i, 2n, 2o. All new compounds were characterized by
1H and 13C NMR spectroscopy and elemental analysis.
Synthesis of 2-Aryl 5-(4-Oxo-3-phenethyl-2-thioxothiazolidiny-
lidenemethyl)furans 11. Knoevenagel condensation26-28 of
N-substituted rhodanine 6 with aldehydes 8a-o in ethanol in
thepresenceofacatalyticamount of 2,2,6,6-tetramethylpiperi-
dine at 78 ꢀC for 1.5-12.5 h gave 11a-o (38-76%) (Scheme 7,
Table 2). Novel 2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazoli-
dinylidenemethyl)furans 11a-o were characterized by 1H and
13C NMR spectroscopy and elemental analysis.
Synthesis of 3-Phenethyl-2-thioxothiazolidin-4-one 6. 3-
Phenethyl-2-thioxothiazolidin-4-one 6 was synthesized by
modification of a literature procedure.21 Phenylethylamine 5
Scheme 3. Synthesis of 2-Bromo-5-iodonicotinic Acid 2na
a Reagents and conditions: (a) i-PrMgCl, I2, THF, room temp, 24 h,
55%; (b) KMnO4, H2O, reflux, 48 h, 14%.
Scheme 4. Synthesis of 4-Methyl-5-iodonicotinic Acid 2oa
Conclusion. In conclusion, we have designed and synthesized
a series of novel 2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazolidi-
nylidenemethyl)furans 11a-o by Suzuki-Miyaura cross-coup-
ling, followed by Knoevenagel condensation. These molecules
Table 1. Preparation of 2-Aryl 5-Formylfurans 8
X of 2, 8, R of 2, 8, R1 of 2, 8, reaction yield (%)
product 8
and 10
and 10
and 10
time, h
of 8a
8a
8b
8c
8d
8e
8f
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
N
H
H
1.5
8.0
89
60
68
51b
59
67
52
80
47
49
74
40
28
31
56
CH3
C2H5
Cl
H
a Reagents and conditions: (a) LDA, MeI, -78 ꢀC, 1 h, 76%; (b) i-Pr-
MgCl, I2, THF, room temp, 24 h, 45%; (c) 5 N HCl, reflux, 24 h, 84%.
H
7.0
H
15
F
H
1.5
7.0
Scheme 5. Synthesis of 3-Phenethyl-2-thioxothiazolidin-4-one 6a
OH
OCH3
H
H
8g
8h
8i
H
3.5
1.0
CH3
H
Br
H
16.0
16.0
7.5
8j
Cl
F
8k
8l
H
H
Cl
H
28.0
28.0
16.0
16.0
8m
8n
8o
N
H
N
Br
H
H
CH3
a Reagents and conditions:(a) (i) CS2, Et2O, 0-5 ꢀC, 0.5 h; (ii) ClCH2-
COOH, EtOH, 1 h, reflux, 56%.
N
a Isolated yields. b Crude yield and used without purification.
Scheme 6. Synthesis of 2-Aryl 5-Formylfurans 8a,b
a Reagents and conditions: (a) (PPh3)2PdCl2, Na2CO3 (aq), EtOH-DME (1:1), 50 ꢀC, 28-80%; (b) Pd(OAc)2, Na2CO3, H2O, room temp, 59-89%.
b For designation of X, R and R1 in 2, 8 and 10 see Table 1.

Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23 7635
Scheme 7. Synthesis of 2-Aryl 5-(4-Oxo-3-phenethyl-2-thioxothiazolidinylidenemethyl)furans 11a
a Reagents and conditions: (a) ethanol, cat. 2,2,6,6-tetramethylpiperidine, reflux, 38-76%. For designation of X, R, and R1 in 8 and 11, see Tables 1
and 2.
Table 2. Preparation of 2-Aryl 5-(4-Oxo-3-phenethyl-2-thioxothiazoli-
dinylidenemethyl)furans 11a-o
using a Whelk-O1 column with detection at 254 nm, a flow rate
of 1.0 mL/min, and methanol as the eluting solvent. Column
chromatography was performed on silica gel 200-425 mesh.
General Procedure for Preparation of 3-(5-Formylfuran-2-
yl)benzoic Acids (8a and 8e). 5-Bromofuran-2-carbaldehyde 9
(0.44 g, 2.5 mmol), sodium carbonate (0.79 g, 7.5 mmol), and
3-carboxyphenylboronic acid 10 (2.75 mmol) were suspended in
degassed water (12.5 mL) under nitrogen. Palladium acetate
(5.6 mg, 0.025 mmol) was added to this mixture, which was
stirred at room temperature for 1.5 h. The slurry was dissolved in
water (100 mL) and filtered through Celite. The filtrate was
acidified with HCl and the precipitate was filtered and washed
with water to give the corresponding 5-(5-formylfuran-2-
yl)benzoic acids 8a and 8e. Analytical samples were obtained
by recrystallization from alcohol.
3-(5-Formylfuran-2-yl)benzoic Acid (8a). Gray microcrystals,
mp 264-266 ꢀC (2-propanol) (lit.30 mp 266-267 ꢀC); 1H NMR
(DMSO-d6) δ 9.63 (s, 1H), 8.37 (br s, 1H), 8.12 (d, J = 8.0 Hz,
1H), 7.99 (d, J = 7.9 Hz, 1H), 7.68-7.62 (m, 2H), 7.41 (d, J =
3.7 Hz, 1H); 13C NMR (DMSO-d6) δ 178.3, 166.9, 157.2, 152.0,
131.9, 130.3, 129.8, 129.3, 129.1, 125.4, 109.7, 108.3. Anal. Calcd
for C12H8O4: C, 66.67; H, 3.73. Found: C, 66.40; H, 3.65.
2-Fluoro-5-(5-formylfuran-2-yl)benzoic Acid (8e). White mi-
crocrystals, mp 231-233 ꢀC (ethanol); 1H NMR (DMSO-d6) δ
9.64 (s, 1H), 8.31 (dd, J = 6.9, 2.4 Hz, 1H), 8.17-8.12 (m, 1H),
7.68 (d, J = 3.7 Hz, 1H), 7.50 (dd, J = 10.5, 8.8 Hz, 1H), 7.40 (d,
J = 3.7 Hz, 1H); 13C NMR (DMSO-d6) δ 178.1, 164.5 (d, J =
3.4 Hz, 1C), 161.5 (d, J = 261.1 Hz, 1C), 156.3, 151.9, 131.2, (d,
J = 9.7 Hz, 1C), 128.2, 125.3 (d, J = 4.0 Hz, 1C), 120.3 (d, J =
11.5 Hz, 1C), 118.3 (d, J = 23.5 Hz, 1C), 109.4. Anal. Calcd for
C12H7FO4: C, 61.54; H, 3.01. Found: C, 61.35; H, 3.24.
reaction
time, h
yield (%)
of 11a
product 11
X of 11
R of 11
R1 of 11
11a
11b
11c
11d
11e
11f
11g
11h
11i
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
N
H
H
1.5
1.5
52
57
48
60
71
50
56
38
54
58
65
48
49
74b
76
CH3
C2H5
Cl
H
H
3.0
3.0
H
F
H
1.5
OH
OCH3
H
H
12.5
2.0
H
CH3
H
1.0
6.0
Br
H
11j
Cl
F
9.0
5.0
11k
11l
H
H
Cl
H
2.0
3.0
11m
11n
11o
N
H
N
Br
H
H
CH3
10.0
10.0
N
a Isolated yield. b One equivalent of 2,2,6,6-tetramethylpiperidine
was used.
exhibited highly potent anti-HIV-1 activity. The most promising
compounds are 11a, 11b, and 11d, which have the lowest EC50
(70-74 nM) and which inhibited replication by laboratory-
adapted and primary HIV-1 strains at <100 nM with selectivity
indexes ranging from 330 to 440, suggesting that these com-
pounds can be used as a lead for developing novel small molecule
HIV fusion/entry inhibitors for treatment of HIV/AIDS patients
who have failed to respond to current antiretroviral therapeutics.
General Procedure for the Preparation of 3-(5-Formylfuran-2-
yl)benzoic and -nicotinic Acids (8b-d,f-o). A mixture of aryl-
(heteryl) bromides/iodides 2b-d,f-o (2.3 mmol), 5-formyl-
2-furanboronic acid 7 (0.42 g, 3.0 mmol), and (Ph3P)2PdCl2
(84.2 mg, 0.12 mmol) in DME (7.0 mL), ethanol (7.0 mL), and 2
M aqueous Na2CO3 (6.9 mL, 13.8 mmol of Na2CO3) was
flushed with nitrogen for 5 min and heated at 50 ꢀC for
3.5-28 h (reaction time is given in Table 1) under nitrogen
atmosphere. The solvents were removed under reduced pres-
sure, the residue was dissolved in water (20 mL), the mixture
obtained was filtered through Celite, and the filtrate was
neutralized with 2 N hydrochloric acid. The solids were filtered,
washed with water, dried, and recrystallized from ethanol to give
3-(5-formylfuran-2-yl)benzoic and -nicotinic acids 8b-d,f-o.
5-(5-Formylfuran-2-yl)-2-methylbenzoic acid (8b). Brownish
microcrystal, mp 236-238 ꢀC (ethanol); 1H NMR (DMSO-d6) δ
9.63 (s, 1H), 8.28 (d, J = 1.8 Hz, 1H), 7.95 (dd, J = 7.8, 1.8 Hz,
1H), 7.67 (d, J = 3.7 Hz, 1H), 7.46 (d, J = 8.1 Hz, 1H), 7.35 (d,
J = 3.7 Hz, 1H), 2.58 (s, 3H); 13C NMR (DMSO-d6) δ 177.9,
168.1, 157.3, 151.7, 140.7, 132.6, 131.4, 128.0, 126.5, 126.3,
125.4, 108.9, 21.2. Anal. Calcd for C13H10O4: C, 67.82; H,
4.38. Found: C, 67.60; H, 4.47.
Experimental Section
Chemistry. Aryl(heteryl) bromides/iodides 2b, 2d-i, 2k-m, 9
and arylboronic acids 10a, 10e were obtained from commercial
suppliers and used without further purification. Aryl(heteryl)
bromides/iodides 2c, 2j, 2n, 2o and 3-phenethyl-2-thioxothiazo-
lidin-4-one 6 were synthesized, and complete details of the
synthesis and spectral data are given in the Supporting Informa-
tion. 5-Formyl-2-furanboronic acid 7 was synthesized according
to a literature method.29 Melting points were determined using a
capillary melting point apparatus equipped with a digital ther-
mometer and are uncorrected. 1H (300 MHz) and 13C (75 MHz)
NMR spectra were recorded in DMSO-d6 (with tetramethylsi-
lanes as the internal standard), unless otherwise stated. Data are
reported as follows: chemical shift, multiplicity (s = singlet, d =
doublet, t = triplet, q = quartet, br s = broad, m = multiplet),
coupling constants (J values) in Hz. Elemental analyses were
performed on a Carlo Erba EA-1108 instrument. All the reac-
tions were performed in flame-dried glassware. The solvents
(ethylene glycol dimethyl ether (DME) and THF) were dried by
the usual methods and distilled before use. Purity of compounds
was determined by elemental analyses and/or HPLC; purity of
target compounds was g95% unless otherwise noted. Analyti-
cal HPLC analyses were performed on Shimadzu SPD-20-A
5-(5-Formylfuran-2-yl)-2-ethylbenzoic Acid (8c). Brown
1
prisms, mp 161-163 ꢀC; H NMR (DMSO-d6) δ 9.63 (s, 1H),

7636 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23
Katritzky et al.
Table 3. Anti-HIV-1 Activity and Selectivity Indexes of the 2-Aryl 5-(4-Oxo-3-phenethyl-2-thioxothiazolidinylidenemethyl)furans 11a-oa
EC50 (μM)
94UG103d
CC50 (μM)
SIe
b
c
product
MW
HIV-1IIIB
HIV-1IIIB
11a
11b
11c
11d
11e
11f
11g
11h
11i
435.5
449.5
463.6
470.0
453.5
451.5
465.6
436.5
514.4
470.0
453.5
449.6
471.0
515.4
450.5
0.044 ( 0.005
0.099 ( 0.010
0.081 ( 0.011
0.051 ( 0.013
0.071 ( 0.010
1.280 ( 0.240
1.300 ( 0.151
0.043 ( 0.008
0.317 ( 0.071
0.064 ( 0.005
0.094 ( 0.002
0.042 ( 0.019
0.232 ( 0.020
0.268 ( 0.055
0.390 ( 0.070
0.098 ( 0.033
0.031 ( 0.016
0.030 ( 0.032
0.017 ( 0.003
0.291 ( 0.241
0.261 ( 0.098
0.266 ( 0.033
0.092 ( 0.004
0.158 ( 0.046
0.078 ( 0.017
0.059 ( 0.014
0.071 ( 0.004
1.010 ( 0.310
0.227 ( 0.080
0.538 ( 0.045
0.074 ( 0.015
0.070 ( 0.012
0.177 ( 0.061
0.073 ( 0.063
0.246 ( 0.037
2.660 ( 0.610
2.229 ( 0.176
0.648 ( 0.108
0.165 ( 0.019
0.033 ( 0.010
0.801 ( 0.138
0.442 ( 0.039
0.598 ( 0.155
0.353 ( 0.009
4.043 ( 0.079
19.35 ( 0.75
42.14 ( 1.61
17.39 ( 0.90
16.82 ( 1.03
27.01 ( 1.48
14.78 ( 1.99
28.72 ( 1.70
26.84 ( 3.73
3.29 ( 1.05
6.73 ( 0.56
78.29 ( 0.57
38.43 ( 2.22
38.37 ( 2.71
5.53 ( 0.08
78.92 ( 4.66
440
426
215
330
380
12
22
624
10
11j
105
835
915
165
40
11k
11l
11m
11n
11o
202
a Each compound was tested in triplicate; the data are presented as the mean ( SD. b The experiment was performed without exclusion of light. c The
experiment was performed with exclusion of light. d 94UG103 is a primary HIV-1 isolate (X4R5, clade A). e SI was calculated on the basis of the CC50 for
MT-2 cells and EC50 for inhibiting infection of HIV-1IIIB
.
8.23 (d, J = 1.9 Hz, 1H), 7.97 (dd, J = 8.0, 1.9 Hz, 1H), 7.67
(d, J=3.7 Hz, 1H), 7.48 (d, J=8.1 Hz, 1H), 7.35 (d, J=3.7 Hz,
1H), 2.97 (q, J=7.4 Hz, 2H), 1.19 (t, J=7.4 Hz, 3H); 13C NMR
(DMSO-d6) δ 177.9, 168.3, 157.4, 151.7, 146.4, 131.4, 131.2,
128.0, 126.4, 126.3, 125.5, 109.0, 26.7, 15.8. Anal. Calcd for
C14H12O4: C, 68.85; H, 4.95. Found: C, 68.51; H, 5.04.
Hz, 1C), 131.0 (d, J=2.9 Hz, 1C), 125.1 (d, J=4.6 Hz, 1C), 125.0
(br s, 1C), 120.8 (d, J=10.3 Hz, 1C), 118.1 (d, J=12.0 Hz, 1C),
113.3 (d, J=12.0 Hz, 1C). Anal. Calcd for C12H7FO4: C, 61.54;
H, 3.01. Found: C, 61.53; H, 2.99.
3-(5-Formylfuran-2-yl)nicotinic Acid (8l). Orange microcrys-
tals, mp 287-289 ꢀC; 1H NMR (DMSO-d6) δ 9.69 (s, 1H), 9.31
(d, J=2.1 Hz, 1H), 9.08 (d, J=1.8 Hz, 1H), 8.62 (t, J=2.1 Hz,
1H), 7.73 (d, J=3.9 Hz, 1H), 7.61 (d, J=3.9 Hz, 1H); 13C NMR
(DMSO-d6) δ 178.5, 165.9, 154.5, 152.6, 150.5, 149.7, 132.5,
127.1, 125.1, 125.0, 111.1. Anal. Calcd for C11H7NO4: C, 60.83;
H, 3.25; N, 6.45. Found: C, 60.47; H, 3.37; N, 6.14.
2-Chloro-5-(5-formylfuran-2-yl)nicotinic Acid (8m). Orange
microcrystals, mp 215-217 ꢀC; NMR (DMSO-d6) δ 9.68 (s,
1H), 9.01 (d, J=2.5 Hz, 1H), 8.56 (d, J=2.3 Hz, 1H), 7.71 (d,
J=3.8 Hz, 1H), 7.58 (d, J=3.7 Hz, 1H); 13C NMR (DMSO-d6)
δ 178.6, 165.5, 153.7, 152.6, 147.9, 147.4, 135.4, 129.5, 125.0,
124.5, 111.5. Anal. Calcd for C11H6ClNO4: C, 52.51; H, 2.40; N,
5.57. Found: C, 52.12; H, 2.58; N, 4.97.
5-(5-Formylfuran-2-yl)-2-hydroxybenzoic Acid (8f). Brown
1
microcrystals, mp 245-247 ꢀC; H NMR (DMSO-d6) δ 9.59
(s, 1H), 8.24 (d, J=2.2 Hz, 1H), 8.03 (dd, J=8.8, 2.3 Hz, 1H),
7.65 (d, J=3.7 Hz, 1H), 7.23 (d, J=3.8 Hz, 1H), 7.11 (d, J=8.8
Hz, 1H); 13C NMR (DMSO-d6) δ 177.5, 171.3, 162. 0, 157.6,
151.4, 132.3, 126.7, 125.8, 120.1, 118.3, 113.8, 107.7. Anal. Calcd
for C12H8O5: C, 62.08; H, 3.47. Found: C, 61.67; H, 3.49.
5-(5-Formylfuran-2-yl)-2-methoxybenzoic Acid (8g). Yellow-
ish microcrystals, mp 220-222 ꢀC; 1H NMR (DMSO-d6) δ 9.59
(s, 1H), 8.12 (d, J=2.3 Hz, 1H), 8.02 (dd, J=8.8, 2.3 Hz, 1H),
7.65 (d, J=3.9 Hz, 1H), 7.30-7.26 (m, 2H), 3.90 (s, 3H); 13C
NMR (DMSO-d6) δ 177.6, 166.8, 159.0, 157.6, 151.4, 129.7,
127.3, 125.8, 122.1, 120.8, 113.3, 107.9, 56.1. Anal. Calcd for
C13H10O5: C, 63.42; H, 4.09. Found: C, 63.22; H, 4.04.
2-Bromo-5-(5-formylfuran-2-yl)nicotinic Acid (8n). Yellow
1
microcrystals; mp 200-210 ꢀC (dec); H NMR ((CD3)2CO) δ
3-(5-Formylfuran-2-yl)-2-methylbenzoic Acid (8h). Yellow
9.81 (s, 1H), 9.07 (d, J = 2.4 Hz, 1H), 8.65 (d, J=2.4 Hz, 1H),
7.67 (d, J = 3.6 Hz, 1H), 7.55 (d, J = 3.6 Hz, 1H); 13NMR
((CD3)2CO) δ 178.5, 165.9, 154.6, 154.2, 148.7, 140.5, 135.9,
131.6, 125.8, 124.2, 111.7. Anal. Calcd for C11H6BrNO4: C,
44.62; H, 2.04; N, 4.73. Found: C, 45.31; H, 2.23; N, 4.24.
4-Methyl-5-(5-formylfuran-2-yl)nicotinic Acid Hemihydrate
(8o). Yellow microcrystals; mp 180 ꢀC (dec); 1H NMR
((CD3)2CO) δ 9.76 (s, 1H), 9.03 (s, 1H), 8.96 (s, 1H), 7.62 (d,
J=3.6 Hz, 1H), 7.15 (d, J=3.9 Hz, 1H), 2.75 (s, 3H); 13NMR
((CD3)2CO) δ 178.5, 167.6, 156.0, 154.1, 152.4, 152.0, 147.1, 128.7,
1
microcrystals, mp 140-142 ꢀC; H NMR (DMSO-d6) δ 9.71
(s, 1H), 8.01 (d, J = 7.8 Hz, 1H), 7.89 (d, J = 7.8 Hz, 1H),
7.42-7.36 (m, 2H), 6.75 (d, J=3.6 Hz, 1H), 2.74 (s, 3H); 13C
NMR (DMSO-d6) δ 177.1, 169.9, 158.3, 151.5, 136.7, 133.1,
131.4, 130.9, 130.1, 125.3, 112.0, 18.3. Anal. Calcd for
C13H10O4: C, 67.82; H, 4.38. Found: C, 67.54; H, 4.32.
5-(5-Formylfuran-2-yl)-2-bromobenzoic Acid Hemihydrate
(8i). Brown microcrystals, mp 161 ꢀC (dec); 1H NMR
(DMSO-d6) δ 9.65 (s, 1H), 8.18 (d, J=2.1 Hz, 1H), 7.90-7.87
(m, 2H), 7.68 (d, J=3.6 Hz, 1H), 7.46 (d, J=3.6 Hz, 1H); 13
C
127.6, 123.6, 114.4, 18.1. Anal. Calcd for C12H9NO4 1/2H2O: C,
3
NMR (DMSO-d6) δ 178.4, 167.0, 156.2, 152.1, 134.9, 128.4,
128.2, 126.6, 125.4, 121.1, 110.3. Anal. Calcd for C12H7-
60.00; H, 4.20; N, 5.83. Found: C, 60.07; H, 3.86; N, 5.26.
General Procedure for Preparation of 2-Aryl 5-(4-Oxo-3-
phenethyl-2-thioxothiazolidinylidenemethyl)furans (11a-o). A
mixture of 3-(5-formylfuran-2-yl)benzoic acid 8 (0.75 mmol),
3-phenethyl-2-thioxothiazolidin-4-one 6 (0.18 g, 0.75 mmol),
and 2 drops of 2,2,6,6-tetramethylpiperidine in 7 mL of ethanol
was heated under reflux for 2.0-12.0 h (reaction time is given in
Table 2). After the mixture was cooled to 40-45 ꢀC, the solid
obtained was filtered and washed with cooled ethanol to give the
corresponding 2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazolidi-
nylidenemethyl)furans (11a-o).
BrO4 1/2H2O: C, 47.40; H, 2.65. Found: C, 47.17; H, 2.52.
3-(5-Formylfuran-2-yl)-2-chlorobenzoic Acid (8j). Yellowish
3
1
microcrystals, mp 192-195 ꢀC; H NMR (DMSO-d6) δ 9.69
(s, 1H), 8.00 (dd, J=1.5, 7.8 Hz, 1H), 7.76 (dd, J=1.5, 7.5 Hz,
1H), 7.70 (d, J=3.9 Hz, 1H), 7.59 (t, J=7.8 Hz, 1H), 7.40 (d, J=
3.6 Hz, 1H); 13C NMR (DMSO-d6) δ 178.6, 167.3, 154.1, 151.9,
135.3, 131.3, 130.5, 128.7, 128.1, 127.9, 124.5, 114.3. Anal. Calcd
for C12H7ClO4: C, 57.51; H, 2.82. Found: C, 57.51; H, 2.85.
3-(5-Formylfuran-2-yl)-2-fluorobenzoic Acid (8k). Orange
1
microcrystals, mp 260-262 ꢀC; H NMR (DMSO-d6) δ 9.69
3-{5-[4-Oxo-3-phenethyl-2-thioxothiazolidin-5-ylidenemethyl]-
furan-2-yl}benzoic Acid (11a). Yellow needles, mp 263-265 ꢀC;
1H NMR (DMSO-d6) δ 8.38 (s, 1H), 8.05 (d, J=7.6 Hz, 1H),
7.97 (d, J=7.6 Hz, 1H), 7.68 (t, J=7.7 Hz, 1H), 7.63 (s, 1H), 7.44
(d, J=3.2 Hz, 1H), 7.36 (d, J=3.3 Hz, 1H), 7.30-7.23 (m, 5H),
(s, 1H), 8.15-8.10 (m, 1H), 7.96-7.91 (m, 1H), 7.72 (d, J=3.7
Hz, 1H), 7.47 (t, J=7.8 Hz, 1H), 7.22 (t, J=3.8 Hz, 1H); 13C
NMR (DMSO-d6) δ 178.3, 164.6 (d, J=2.9 Hz, 1C), 157.7 (d, J=
264.5 Hz, 1C), 151.6, 151.6 (d, J=2.3 Hz, 1C), 132.7 (d, J=1.4

Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23 7637
4.22 (t, J = 7.1 Hz, 2H), 2.95 (t, J = 7.0 Hz, 2H); 13C NMR
(DMSO-d6) δ 193.7, 174.5, 166.7, 166.3, 156.9, 149.5, 137.6,
131.9, 129.8, 128.9, 128.7, 128.5, 128.4, 126.6, 124.9, 123.0,
118.9, 118.2, 111.0, 45.2, 32.1. Anal. Calcd for C23H17NO4S2:
C, 63.43; H, 3.93; N, 3.22. Found: C, 63.21; H, 3.80; N, 3.20.
2-Methyl-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}benzoic Acid (11b). Yellow microcrystals,
7.42 (d, J=3.6 Hz, 1H), 7.31-7.24 (m, 5H), 7.13 (d, J=3.6 Hz,
1H), 4.24 (t, J=7.8 Hz, 2H), 2.96 (t, J=8.2 Hz, 2H), 2.61 (s, 3H);
13C NMR (DMSO-d6) δ 193.7, 166.3, 157.3, 149.2, 137.6, 135.0,
130.4, 130.0, 128.7, 128.5, 126.6, 126.3, 122.7, 118.8, 118.4,
114.5, 45.3, 32.1, 18.5. Anal. Calcd for C24H19NO4S2: C,
64.12; H, 4.26; N, 3.12. Found: C, 63.75; H, 4.42; N, 3.14.
2-Bromo-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}benzoic Acid (11i). Deep-red microcrystals;
mp 241 ꢀC (dec); 1H NMR (DMSO-d6) δ 8.20 (d, J=2.1 Hz, 1H),
7. 94-7.98 (m, 1H), 7.86-7.89 (m, 1H), 7.69 (s, 1H), 7.54 (d, J=
3.6 Hz, 1H), 7.41 (d, J=3.6 Hz, 1H), 7.24-7.38 (m, 6H), 4.27 (t,
J=8.4 Hz, 2H), 3.00 (t, J=7.5 Hz, 2H); 13C NMR (DMSO-d6) δ
193.7, 167.1, 166.4, 156.0, 149.8, 137.7, 135.1, 135.0, 128.8,
128.6, 128.0, 127.5, 126.7, 126.0, 123.1, 120.4, 119.3, 118.2,
111.7, 45.4, 32.2. HRMS (DIP-CI-MS) calcd for C23H16-
BrNO4S2: [M þ H] 513.9779. Found: 513.9782. Anal. HPLC:
98.8%, tR=3.38 (tracing, 0.7%, tR=4.19).
2-Chloro-3-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}benzoic Acid (11j). Orange microcrystals,
mp 258-260 ꢀC; 1H NMR (DMSO-d6) δ 8.0 (dd, J=7.8, 1.5 Hz,
1H), 7.75-7.64 (m, 3H), 7.47 (d, J=3.6 Hz, 1H), 7.40 (d, J=
3.6 Hz, 1H), 7.33-7.29 (m, 2H), 7.24-7.21 (m, 3H), 4.23 (t, J=
7.5 Hz, 2H), 2.96 (t, J=7.5 Hz, 2H); 13C NMR (DMSO-d6) δ
193.9, 167.4, 166.5, 154.0, 149.7, 137.8, 135.7, 130.3, 130.0,
128.9, 128.7, 128.5, 128.1, 127.4, 126.8, 122.5, 120.0, 118.3,
115.9, 45.5, 32.3. Anal. Calcd for C23H16ClNO4S2: C, 58.78;
H, 3.43; N, 2.98. Found: C, 58.40; H, 3.39; N, 2.88.
1
mp 297-298 ꢀC; H NMR (DMSO-d6) δ 8.27 (d, J=1.8 Hz,
1H), 7.88 (dd, J=7.8, 1.8 Hz, 1H), 7.63 (s, 1H), 7.49 (d, J=
8.1 Hz, 1H), 7.34-7.21 (m, 7H), 4.23 (t, J=7.7 Hz, 2H), 2.96 (t,
J=7.6 Hz, 2H), 2.57 (s, 3H); 13C NMR (DMSO-d6) δ 193.7,
168.1, 166.3, 157.2, 149.2, 140.2, 137.6, 132.7, 131.5, 128.7,
128.5, 127.2, 126.6, 126.4, 125.9, 123.2, 118.5, 118.3, 110.4,
45.3, 32.2, 21.2. Anal. Calcd for C24H19NO4S2: C, 64.12; H,
4.26; N, 3.12. Found: C, 63.85; H, 4.21; N, 3.02.
2-Ethyl-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylidene-
methyl]furan-2-yl}benzoic Acid (11c). Orange microcrystals, mp
251-253 ꢀC; 1H NMR (DMSO-d6) δ 8.22 (d, J=1.92 Hz, 1H),
7.91 (dd, J=8.1, 1.92 Hz, 1H), 7.63 (s, 1H), 7.51 (d, J=8.1 Hz,
1H), 7.36-7.29 (m, 4H), 7.24-7.21 (m, 3H), 4.22 (t, J=7.4 Hz,
2H), 3.00-2.93 (m, 4H), 1.20 (t, J = 7.6 Hz, 3H); 13C NMR
(DMSO-d6) δ 193.7, 168.3, 166.3, 157.2, 149.3, 145.9, 137.6,
131.5, 131.3, 128.7, 128.5, 127.2, 126.6, 126.3, 125.9, 123.2,
118.5, 118.3, 110.4, 45.2, 32.2, 26.7, 15.8. Anal. Calcd for
C25H21NO4S2: C, 64.77; H, 4.57; N, 3.02. Found: C, 64.66; H,
4.48; N, 3.04.
2-Chloro-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}benzoic Acid (11d). Orange microcrystals,
2-Fluoro-3-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}benzoic Acid (11k). Orange microcrystals,
1
mp 276-278 ꢀC; H NMR (DMSO-d6) δ 8.21 (d, J=2.2 Hz,
1
1H), 7.93 (dd, J=8.5, 2.2 Hz, 1H), 7.75 (d, J=8.5 Hz, 1H), 7.64
(s, 1H), 7.48 (d, J = 3.7 Hz, 1H), 7.37 (d, J = 3.8 Hz, 1H),
7.33-7.29 (m, 2H), 7.25-7.21 (m, 3H), 4.23 (t, J=7.1 Hz, 2H),
2.96 (t, J=7.1 Hz, 2H); 13C NMR (DMSO-d6) δ 193.6, 166.3,
166.3, 155.9, 149.8, 137.6, 132.6, 131.9, 131.8, 128.7, 128.6,
127.6, 127.5, 126.7, 126.3, 123.0, 119.2, 118.1, 111.6, 45.3,
32.2. Anal. Calcd for C23H16ClNO4S2: C, 58.78; H, 3.43; N,
2.98. Found: C, 59.04; H, 3.28; N, 2.95.
mp 262-264 ꢀC; H NMR (DMSO-d6) δ 8.00-7.95 (m, 1H),
7.87-7.81 (m, 1H), 7.65 (s, 1H), 7.50 (t, J=7.7 Hz, 1H), 7.39 (d,
J=3.7 Hz, 1H), 7.33-7.29 (m, 2H), 7.25-7.18 (m, 4H), 4.22 (t,
J=8.0 Hz, 2H), 2.95 (t, J=8.0 Hz, 2H); 13C NMR (DMSO-d6)
δ 193.7, 166.3, 164.9, 157.2 (d, J=263.4 Hz, 1C), 151.6, 149.3,
137.6, 132.0, 129.4 (br s, 1C), 128.7, 128.6, 126.7, 125.1 (d, J=
4.6 Hz, 1C), 122.9, 122.2 (br d, J=10.9 Hz, 1H), 119.6, 118.0,
117.9 (d, J=14.9 Hz, 1C), 114.6 (d, J=13.2 Hz, 1C), 45.3, 32.1.
Anal. Calcd for C23H16FNO4S2: C, 60.91; H, 3.56; N, 3.09.
Found: C, 61.13; H, 3.52; N, 3.11.
2-Fluoro-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}benzoic Acid (11e). Yellow microcrystals,
mp 286-288 ꢀC; 1H NMR (DMSO-d6) δ 8.29 (dd, J=6.7, 2.2
Hz, 1H), 8.07-8.02 (m, 1H), 7.62 (s, 1H), 7.53 (dd, J=10.4, 8.9
Hz, 1H), 7.40 (d, J = 3.8 Hz, 1H), 7.35 (d, J = 3.8 Hz, 1H),
7.31-7.21 (m, 5H), 4.22 (t, J=7.6 Hz, 2H), 2.95 (t, J=7.6 Hz,
2H); 13C NMR (DMSO-d6) δ 193.6, 166.3, 164.4 (d, J=3.4 Hz,
1C), 161.1 (d, J=260.5 Hz, 1C), 156.1, 149.5, 137.6, 130.2 (d, J=
9.2 Hz, 1C), 128.7, 128.5, 127.7, 126.6, 125.2 (d, J=3.4 Hz, 1C),
123.1, 120.3 (d, J=11.5 Hz, 1C), 118.8, 118.4 (d, J=23.5 Hz,1C),
118.2, 110.8, 45.2, 32.1. Anal. Calcd for C23H16FNO4S2: C,
60.91; H, 3.56; N, 3.09. Found: C, 60.90; H, 3.38; N, 3.06.
2-Hydroxy-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-yli-
denemethyl]furan-2-yl}benzoic Acid (11f). Orange microcrystals,
5-{5-[4-Oxo-3-phenethyl-2-thioxothiazolidin-5-ylidenemethyl]-
furan-2-yl}nicotinic Acid (11l). Orange microcrystals, mp
259-261 ꢀC; 1H NMR (DMSO-d6) δ 9.14 (d, J=2.2 Hz, 1H),
9.03 (d, J=1.9 Hz, 1H), 8.58 (t, J=2.1 Hz, 1H), 7.67 (s, 1H),
7.56 (d, J=3.7 Hz, 1H), 7.40 (d, J=3.7 Hz, 1H), 7.34-7.22 (m,
5H), 4.24 (t, J=7.7 Hz, 2H), 2.97 (t, J=7.7 Hz, 2H); 13C NMR
(DMSO-d6) δ 193.6, 166.3, 166.2, 154.9, 150.2, 150.0, 147.3,
137.6, 131.6, 130.9, 128.7, 128.5, 126.6, 124.4, 122.8, 119.4,
118.1, 111.8, 45.3, 32.1. Anal. Calcd for C22H16N2O4S2: C,
60.54; H, 3.69; N, 6.42. Found: C, 60.44; H, 3.65; N, 6.28.
2-Chloro-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}nicotinic Acid (11m). Orange microcrystals,
1
1
mp 257-259 ꢀC; H NMR (DMSO-d6) δ 8.24 (d, J=2.2 Hz,
mp 246-248 ꢀC; H NMR (DMSO-d6) δ 8.71 (d, J=2.3 Hz,
1H), 8.00 (dd, J=8.7, 2.2 Hz, 1H), 7.59 (s, 1H), 7.34-7.21 (m,
7H), 7.14 (d, J=8.7 Hz, 1H), 4.22 (t, J=7.3 Hz, 2H), 2.95 (t, J=
7.3 Hz, 2H). Anal. Calcd for C23H17NO5S2: C, 61.18; H, 3.79; N,
3.10. Found: C, 61.02; H, 3.75; N, 3.09.
1H), 8.12 (d, J=2.5 Hz, 1H), 7.66 (s, 1H), 7.51 (d, J=3.9 Hz,
1H), 7.38 (d, J=3.7 Hz, 1H), 7.33-7.29 (m, 2H), 7.24-7.22
(m, 3H), 4.23 (t, J=7.3 Hz, 2H), 2.96 (t, J=7.3 Hz, 2H); 13C
NMR (DMSO-d6) δ 193.6, 166.6, 166.3, 154.5, 149.9, 146.4,
143.1, 138.3, 137.6, 132.1, 128.7, 128.6, 126.7, 123.8, 122.8,
119.4, 118.1, 11.9, 45.3, 32.2. Anal. Calcd for C22H15ClN2O4S2:
C, 56.11; H, 3.21; N, 5.95. Found: C, 56.01; H, 3.01; N, 5.60.
2-Bromo-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}nicotinic Acid Hemi 2,2,6,6-Tetramethylpi-
peridine Dihydrate (11n). Orange microcrystals; mp 200-
2-Methoxy-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-yli-
denemethyl]furan-2-yl}benzoic Acid (11g). Orange microcrys-
tals, mp 221-223 ꢀC; 1H NMR (DMSO-d6) δ 8.14 (d, J =
2.3 Hz, 1H), 7.99 (dd, J = 8.8, 2.3 Hz, 1H), 7.64 (s, 1H),
7.40-7.31 (m, 5H), 7.28-7.24 (m, 3H), 4.26 (t, J = 7.3 Hz,
2H), 3.93 (s, 3H), 2.98 (t, J=8.0 Hz, 2H); 13C NMR (DMSO-d6)
δ 193.6, 166.8, 166.3, 158.6, 157.5, 148.8, 137.7, 128.9, 128.7,
128.5, 126.8, 126.6, 123.5, 122.3, 120.8, 118.3, 117.7, 113.5,
109.4, 56.1, 45.2, 32.2. Anal. Calcd for C24H19NO5S2: C,
61.92; H, 4.11; N, 3.01. Found: C, 61.68; H, 4.02; N, 3.02.
2-Methyl-3-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylide-
nemethyl]furan-2-yl}benzoic Acid (11h). Red microcrystals,
1
205 ꢀC; H NMR (DMSO-d6) δ 8.79 (d, J=2.4 Hz, 1H), 8.41
(br s, 1H), 8.25 (d, J=2.4 Hz, 1H), 7.64 (s, 1H), 7.55 (d, J=3.9
Hz, 1H), 7.37 (d, J=3.9 Hz, 1H), 7.33-7.21 (m, 5H), 4.22 (t,
J=7.2 Hz, 2H), 2.95 (t, J=7.8 Hz, 2H); 13C NMR (DMSO-d6) δ
193.6, 166.7, 166.3, 153.8, 150.2, 145.3, 138.1, 137.6, 132.6,
128.7, 128.6, 126.7, 124.2, 122.7, 119.8, 118.0, 112.6, 55.6,
45.3. Anal. Calcd for C22H15S2BrN2O4 1/2C9H19N 2H2O: C,
1
mp 248-250 ꢀC; H NMR (DMSO-d6) δ 7.85 (d, J=7.6 Hz,
1H), 7.78 (d, J=7.7 Hz, 1H), 7.69 (s, 1H), 7.50 (t, J=7.8 Hz, 1H),
3
3
52.69; H, 4.42; N, 5.80. Found: C, 52.84; H, 4.02; N, 5.32.

7638 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 23
Katritzky et al.
4-Methyl-5-{5-[4-oxo-3-phenethyl-2-thioxothiazolidin-5-ylidene-
methyl]furan-2-yl}nicotinic Acid (11o). Brown microcrystals; mp
270 ꢀC (dec); 1H NMR (DMSO-d6) δ 8.97 (s, 1H), 8.89 (s, 1H),
7.71 (s, 1H), 7.43 (d, J=3.9 Hz, 1H), 7.34-7.22 (m, 6H), 4.23 (t,
J=6.9 Hz, 2H), 2.96 (t, J=7.2 Hz, 2H), 2.68 (s, 3H); 13C NMR
(DMSO-d6) δ 193.6, 167.6, 166.3, 154.4, 150.2, 150.0, 144.4, 137.7,
128.8, 128.7, 128.6, 126.7, 126.0, 122.3, 119.6, 118.2, 115.5, 45.3,
32.2, 18.1. Anal. Calcd for C23H18S2N2O4: C, 61.32; H, 4.03; N,
6.22. Found: C, 61.06; H, 4.05; N, 5.94.
Molecular Modeling and Docking of 11d onto the gp41 Hydro-
phobic Cavity. We used the automated docking software Glide
(Schrodinger, Portland, OR). This applies a two-stage scoring
process to sort out the best conformations and orientations of
the ligand (defined as pose) based on its interaction pattern with
the receptor. The starting point of the docking simulation was
the X-ray structure of the gp41 core (1AIK) available from the
Protein Data Bank (PDB) at the “Research Collaboratory for
Structural Bioinformatics (RCSB)”; this has been extensively
used in our research program. Three-dimensional coordinates
of the ligands and their isomeric, ionization, and tautomeric
states were generated using the LigPrep (including Ionizer)
module from Schrodinger. The protein was prepared using the
protein preparation tool available in the software. A grid file
encompassing the area in the cavity that contains information
on the properties of the associated receptor was created. Con-
formational flexibility of the ligands was handled via an ex-
haustive conformational search. We used Schrodinger’s
proprietary GlideScore scoring function in standard precision
(SP) and extra precision (XP) mode to score the optimized poses.
The poses were selected on the basis of the salt bridge and other
hydrophobic and hydrogen bond interactions.
added to the virus (HIV-1IIIB) and MT-2 cells in the biosafety
hood with the light turned off. Then the plates were wrapped
with foil before they were put in the CO2 incubator. The other
procedures are the same as described above.
Inhibitory activity of compounds on infection by a primary
HIV-1 isolate, 94UG103 (X4R5, clade A), was determined as
previously described.18 Peripheral blood mononuclear cells
(PBMCs) were isolated from the blood of healthy donors at
the New York Blood Center by standard density gradient
centrifugation using Histopaque-1077 (Sigma). The cells were
plated in 75 cm2 plastic flasks and incubated at 37 ꢀC for 2 h. The
nonadherent cells were collected and resuspended at 5 ꢀ 106 in
10 mL of RPMI-1640 medium containing 10% FBS, 5 μg/mL
PHA, and 100 U/mL IL-2 (Sigma), followed by incubation at
37 ꢀC for 3 days. The PHA-stimulated cells were infected with
94UG103 at 0.01 multiplicity of infection (MOI) in the absence
or presence of a compound at graded concentrations. Culture
media were changed every 3 days. The supernatants were
collected 7 days postinfection and tested for p24 antigen by
ELISA as described above. The percent inhibition of p24
production and EC50 values were calculated as described above.
Assessment of in Vitro Cytotoxicity. The in vitro cytotoxicity
of compounds on MT-2 cells was measured by XTT assay.19
Briefly, an amount of 100 μL of the test compound at graded
concentrations was added to equal volumes of cells (5 ꢀ 105/mL)
in wells of 96-well plates. After incubation at 37 ꢀC for 4 days,
50 μL of XTT solution (1 mg/mL) containing 0.02 μM phena-
zine methosulfate (PMS) was added. After 4 h, the absorbance
at 450 nm was measured with an ELISA reader. The CC50
(concentration for 50% cytotoxicity) values were calculated
using the CalcuSyn program.32
Determination of the Inhibitory Activity of the Compounds on
HIV-1 Replication. The inhibitory activity of compounds on
HIV-1IIIB replication in MT-2 cells was determined as pre-
viously described.18 In brief, 1 ꢀ 104 MT-2 cells were infected
with an HIV-1IIIB strain (100 TCID50) in 200 μL of RPMI 1640
medium containing 10% FBS in the presence or absence of a test
compound at graded concentrations overnight. Then the culture
supernatants were removed and fresh media containing no test
compounds were added. On the fourth day postinfection, an
amount of 100 μL of culture supernatants was collected from
each well, mixed with equal volumes of 5% Triton X-100, and
assayed for p24 antigen, which was quantitated by ELISA.
Briefly, wells of polystyrene plates (Immulon 1B, Dynex Tech-
nology, Chantilly, VA) were coated with HIV immunoglobulin
(HIVIG), which was prepared from plasma of HIV-seropositive
donors with high neutralizing titers against HIV-1IIIB, in
0.085 M carbonate-bicarbonate buffer (pH 9.6) at 4 ꢀC over-
night, followed by washing with buffer (0.01 M PBS containing
0.05% Tween-20) and blocking with PBS containing 1% dry fat-
free milk (Bio-Rad, Inc., Hercules, CA). Virus lysates were
added to the wells and incubated at 37 ꢀC for 1 h. After extensive
washes, anti-p24 mAb (183-12H-5C), biotin-labeled antimouse
IgG1 (Santa Cruz Biotechnolgy, Santa Cruz, CA), streptavidin-
labeled horseradish peroxidase (Zymed, S. San Francisco, CA),
and the substrate 3,30,5,50-tetramethylbenzidine (Sigma Chemi-
cal Co., St. Louis, MO) were added sequentially. Reactions were
terminated by addition of 1 N H2SO4. Absorbance at 450 nm
was recorded in an ELISA reader (Ultra 386, TECAN, Research
Triangle Park, NC). Recombinant protein p24 purchased from
United States Biological (Swampscott, MA) was included for
establishing standard dose-response curves. Each sample was
tested in triplicate. The percentage of inhibition of p24 produc-
tion was calculated as previously described.31 EC50 values were
calculated using the computer program CalcuSyn,32 kindly
provided by Dr. T. C. Chou (Sloan-Kettering Cancer Center,
New York).
Acknowledgment. This study was supported by NIH
Grant RO1 AI46221 and the intramural fund from the New
York Blood Center. We thank Dr. C. D. Hall for helpful
discussions.
Supporting Information Available: Experimental details,
spectral data for aryl(heteryl) bromides/iodides 2c, 2j, 2n, 2o,
intermediates 3n, 3o, 4o, 3-phenethyl-2-thioxothiazolidin-4-one
6, and the inhibition of compounds 11a, 11b, and 11d on
infection by HIV-1IIIB and 94UG103. This material is available
free of charge via the Internet at http://pubs.acs.org.
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