Computer-assisted design, synthesis and biological evaluation of novel pyrrolyl heteroaryl sulfones targeted at HIV-1 reverse transcriptase as non-nucleoside inhibitors

Article abstract of DOI:10.1016/S0968-0896(00)00144-9

Three pyrrolyl heteroaryl sulfones (ethyl 1-[(1H-benzimidazol-2(3H)one-5-yl)sulfonyl]-1H-pyrrole-2-carboxylate, ethyl 1-[(1H-benzimidazol-5(6)-yl)sulfonyl]-1H-pyrrole-2-carboxylate and ethyl 1-[(1H-benzotriazol-5(6)-yl)sulfonyl]-1H-pyrrole-2-carboxylate) were designed as novel HIV-1 reverse transcriptase non-nucleoside inhibitors using structure-based computational methods. Although these compounds were inactive in the cell-based assay, they inhibited the target enzyme with micromolar potency (IC₅₀s=2μM, 3μM and 9μM, respectively). Copyright (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0968-0896(00)00144-9

Bioorganic & Medicinal Chemistry 8 (2000) 2305±2309
Computer-Assisted Design, Synthesis and Biological Evaluation of
Novel Pyrrolyl Heteroaryl Sulfones Targeted at HIV-1 Reverse
Transcriptase as Non-Nucleoside Inhibitors
Romano Silvestri,a Marino Artico,a,* Gabriella De Martino,a Ettore Novellino,b
Giovanni Greco,b Antonio Lavecchia,b Silvio Massa,c Anna Giulia Loi,d
Silvia Doratiottod and Paolo La Collad
a
Á
Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Studi Farmaceutici, Universita degli Studi di Roma
``La Sapienza'', P.le A. Moro 5, Box 36-Roma 62 I-00185 Roma, Italy
b
Á
Dipartimento di Chimica Farmaceutica e Tossicologica, Universita degli Studi di Napoli ``Federico II'',
Via D. Montesano 49, I-80131 Napoli, Italy
c
Á
Dipartimento Farmaco Chimico Tecnologico, Universita degli Studi di Siena, Via A. Moro, I-53100 Siena, Italy
d
Á
Dipartimento di Biologia Sperimentale, Sezione di Microbiologia, Universita degli Studi di Cagliari,
Cittadella Universitaria, SS 554, I-09042 Monserrato (Cagliari), Italy
Received 29 March 2000; accepted 31 May 2000
AbstractÐThree pyrrolyl heteroaryl sulfones (ethyl 1-[(1H-benzimidazol-2(3H)one-5-yl)sulfonyl]-1H-pyrrole-2-carboxylate, ethyl 1-
[(1H-benzimidazol-5(6)-yl)sulfonyl]-1H-pyrrole-2-carboxylate and ethyl 1-[(1H-benzotriazol-5(6)-yl)sulfonyl]-1H-pyrrole-2-carboxylate)
were designed as novel HIV-1 reverse transcriptase non-nucleoside inhibitors using structure-based computational methods.
Although these compounds were inactive in the cell-based assay, they inhibited the target enzyme with micromolar potency
(IC50s=2 mM, 3 mM and 9 mM, respectively). # 2000 Elsevier Science Ltd. All rights reserved.
Introduction
Reverse transcriptase (RT) is a key enzyme in the half life
of the human immuno-de®ciency virus (HIV) which cat-
alyzes the conversion of RNA retroviral genome into
proviral DNA.1,2 RT inhibitors can be classi®ed into two
categories: nucleoside and non-nucleoside compounds,
their prototypes being zidovudine (1)3 and nevirapine
(2),4 respectively, both currently employed in the treat-
ment of AIDS.5 The former are converted intracellularly
into triphosphates derivatives and act as DNA chain-
terminating analogues of the natural deoxy-nucleoside
triphosphates; the latter directly inhibit RT, non-
competitively, upon binding to an allosteric site located
approximately 10 A from the polymerase catalytic site.6
While nucleoside inhibitors block RT of either type-1
and type-2 viruses (HIV-1 and HIV-2), non-nucleoside
compounds inhibit selectively RT from HIV-1.
In 1996 we described pyrrolyl aryl sulfones (PASs) as a
new class of RT non-nucleoside inhibitors.7 The most
potent inhibitor of that series, compound 3 (EC50=
0.14 mM, IC50=0.4 mM and SI>1429), was more
recently taken as a lead for a computer-assisted optimi-
zation project.9 Using the three-dimensional structure of
RT co-crystallized with the a-APA derivative R95845,8
we developed a model of RT/3 complex (Fig. 1) which
guided the design of novel PASs with improved inhibitory
activity.9 Among the new inhibitors synthesized and
tested, compound 4 was the most active with EC50=
0.045 mM, IC50=0.05 mM and SI=5333. Compared
with the lead 3, these values represented a 3- and 8-fold
*Corresponding author. Tel. and fax: +39-06-4462731; e-mail: arti-
co@uniroma1.it
0968-0896/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(00)00144-9

2306
R. Silvestri et al. / Bioorg. Med. Chem. 8 (2000) 2305±2309
Figure 2. Theoretical model of the RT/5 complex. Two hydrogen
bonds are hypothesized to occur betwen the ureidic NH groups of the
docked molecule and the carbonyl oxygens of Lys101 and His235.
Only a subset of residues of the non-nucleoside binding site closest to
the ligand is displayed.
Figure 1. Compound 3 docked into the RT binding site of non-
nucleoside inhibitors. Only a subset of residues closest to the ligand is
displayed for the sake of clarity.
improvement of the in vitro antiviral and RT inhibitory
activity, respectively, together with the highest selectivity
achieved in the PAS series.
6 and 7 were also proposed for synthesis and biological
testing. We reasoned that these compounds might
`choose' the energetically most favorable hydrogen
bond between those feasible with Lys101 and His235
carbonyls by simply switching from one tautomeric
form to another (see Fig. 3).
As a further development of our strucuture-based design
strategy we sought novel leads targeted at the HIV-1 RT
as non-nucleoside inhibitors structurally related to com-
pound 3. This problem was approached by replacing the
pharmacophoric p-chloroaniline moiety, common to all
potent PAS derivatives, with a heteroaromatic system
containing at least one hydrogen bond donor group for
the Lys101 carbonyl. According to our theoretical model
of RT/PAS interaction, a hydrogen bond between the
NH2 of 3 and the Lys101 oxygen (Fig. 1) was in fact
mandatory for high inhibitory activity.9
Chemistry
The synthetic routes for the novel pyrrolyl heteroaryl
sulfones 5, 6 and 7 are shown in Scheme 1. Ethyl 1-[(4-
¯uoro-3-nitrophenyl)sulfonyl]-1H-pyrrole-2-carboxylate
(8) was prepared by adding a suspension of the potassium
salt of ethyl 1H-pyrrole-2-carboxylate10 to a solution of 4-
¯uoro-3-nitrobenzenesulfonyl chloride.11 Treatment of 8
with an excess of aqueous ammonium hydroxide at
room temperature furnished ethyl 1-[(4-amino-3-nitro-
phenyl)sulfonyl]-1H-pyrrole-2-carboxylate (9) which
was reduced to the diamino derivative 10 with powdered
iron in glacial acetic acid. Compound 10 was trans-
formed into ethyl 1-[(1H-benzimidazol-2(3H)one-5-yl)
sulfonyl]-1H-pyrrole-2-carboxylate (5) by reaction with
bis(trichloromethyl) carbonate (triphosgene)12 in the pre-
sence of triethylamine, into ethyl 1-[(1H-benzimidazol-
5(6)-yl)sulfonyl]-1H-pyrrole-2-carboxylate (6) by heating
The docking of the o-phenylenurea derivative 5 (Fig. 2)
brought out the possibility to retain the NHÁ Á ÁO=C
(Lys101) hydrogen bond and to engage an additional
hydrogen bond with the His235 carbonyl. Both hydrogen
bonds were expected to occur via the two ureidic NH
groups. The benzimidazole and benzotriazole derivatives

R. Silvestri et al. / Bioorg. Med. Chem. 8 (2000) 2305±2309
2307
Figure 3. Theoretical model of the RT/6 complex. Two tautomeric forms of the benzimidazole derivative 6 were considered di€ering in the position
of the NH group. This latter donates a hydrogen bond to the carbonyl oxygen of Lys101 (tautomer on the left) or His235 (tautomer on the right).
Only residues Lys101 and His 235 of the non-nucleoside binding site are displayed.
Table 1. Cytotoxicity, anti-HIV-1 activity in MT-4 cells, selectivity
index and RT inhibitory activity valuesa
b
c
e
Compound
CC50
EC50
SId
IC50
5
6
7
160
40
200
>200
>200
21
>40
>2000
7.6
2
3
9
0.4
0.6
Ð
Ð
3
Nevirapine
0.14
0.35
>1429
>571
aData represent mean values for three separate experiments.Variation
among triplicate samples was less than 15%.
bDose (mM) required to reduce the viability of mock-infected cells by
50% as determined by the MTT method.
cDose (mM) required to achieve 50% protection of MT-4 cells from
HIV-1 induced cytopathicity as determined by the MTT method.
dSelectivity index expressed as CC50/EC50 ratio.
eDose (mM) required to inhibit the HIV-1 RT activity by 50%.
RT) values. Data relative to nevirapine and 3 are also
reported as references.
The compounds designed as potential anti-RT new leads
were found devoid of activity in the cell-based assay but
moderately active when tested against the enzyme
(IC50s=2 mM, 3 mM and 9 mM, respectively). The lower
potency of 5±7, compared with 3, might depend on
intramolecular hydrogen bonds stabilizing the RT-bound
conformation of 3 which are absent in 5±7. Particularly,
the docked conformation of 3 features two hydrogen
bonds donated by the NH2 group to sulfone and ester
oxygens (see Fig. 1). Additional reasons for the relatively
low activity of the novel inhibitors might be the missing
chlorine which makes van der Waals interactions in the
RT/3 complex and the partial negative charge of the
non protonated nitrogen of 6 and 7 facing the negative
electrostatic potential of the Lys101 or His235 carbonyl.
In designing the new ligands, it was actually hoped that
the favorable e€ects of the hydrogen bonds attributed
to the virtual RT/inhibitor complexes (Figs 2 and 3)
could counterbalance the above mentioned unfavorable
Scheme 1. Reagents and reaction conditions: (i) t-BuOK 18-C-6,
THF, r.t., 3.5 h; (ii) 30% NH4OH, dioxane, r.t., 30 min.; (iii) Fe,
CH3COOH, 60 ꢀC, 2 h; (iv) CO(OCCl3)2, (C2H5)3N, r.t., 24 h; (v)
HCOOH, 100 ꢀC, 2 h; (vi) NaNO2, CH3COOH, r.t., 1 h.
with formic acid, and into ethyl 1-[(1H-benzotriazol-5(6)-
yl)sulfonyl]-1H-pyrrole-2-carboxylate (7) by reacting with
sodium nitrite in aqueous acetic acid.
Results and Discussion
Table 1 summarizes the results of the biological evalua-
tions of compounds 5±7, expressed as CC50 (cytotoxicity),
EC50 (antiproliferative activity), SI (selectivity index) and
IC50 (inhibitory activity against wild type recombinant

2308
R. Silvestri et al. / Bioorg. Med. Chem. 8 (2000) 2305±2309
factors. To sum up, our structure-based design approach
a€orded novel HIV-1 RT non-nucleoside inhibitors
characterized by micromolar potency.
C, H, N, F, S. Further elution with the same solvent
furnished ethyl 1-{[4-(2-ethoxycarbonyl-1H-pyrrole-1-yl)-
3 - nitrophenyl]sulfonyl} - 1H - pyrrole-2-carboxylate
(0.14 g, 3%); mp 92±94 ꢀC (toluene/cyclohexane). 1H
NMR (CDCl3): d 1.18 and 1.30 (two t, J=7.0Hz, 6H,
COOCH2CH3), 4.11 and 4.23 (two q, J=7.0 Hz, 4H,
COOCH2CH3), 6.35±6.47 (m, 2H, pyrrole), 6.91 (m, 1H,
pyrrole), 7.06±7.18 (m, 2H, pyrrole), 7.59 (d, J=8.3 Hz,
1H, H5-benzene), 7.72 (m, 1H, pyrrole), 8.39 (dd,
J=2.1 and 8.3 Hz, 1H, H6-benzene), 8.64 ppm (d,
J=2.1 Hz, 1H, H2-benzene). IR (Nujol): n 1700 and
1720 cm 1 (CO). Anal. C20H19N3O8S (461.44), C, H, N, S.
Experimental
Chemical procedures
Melting points (mp) were determined on a Buchi 510
apparatus and are uncorrected. Infrared spectra (IR) were
run on a Perkin±Elmer 1310 spectrophotometer and on a
Jasco FT/DR-200 di€use re¯ectance spectrophotometer.
1
Band position and absorption ranges are given in cm
.
Ethyl 1-[(4-amino-3-nitrophenyl)sulfonyl]-1H-pyrrole-2-
carboxylate (9). A solution of 8 (1.00 g, 0.0029 mol) in
dioxane (10 mL) was treated at room temperature under
stirring with 30% ammonium hydroxide (7.7 mL) for
30 min. Water and ethyl acetate were added by shaking.
The organic layer was separated, washed with brine and
dried. Removal of the solvent furnished satisfactory
pure 9 (0.9 g, 91%); mp 129±130 ꢀC (toluene/ligroin). 1H
NMR (CDCl3): d 1.29 (t, J=7.2 Hz, 3H, COO
CH2CH3), 4.19 (q, J=7.2 Hz, 2H, COOCH2CH3), 6.30
(m, 1H, pyrrole), 6.59 (broad s, 2H, NH2, disappeared
on treatment with D2O), 6.88 (d, J=9.1 Hz, 1H, H5-
benzene), 7.04 (m, 1H, pyrrole), 7.68 (m, 1H, pyrrole),
8.00 (dd, J=2.3 and 9.1 Hz, 1H, H6-benzene), 8.79 ppm
(d, J=2.3 Hz, 1H, H2-benzene). IR (Nujol): n 1690
Proton nuclear magnetic resonance (1H NMR) spectra
were recorded on Bruker AM-200 (200 MHz) and Varian
Gemini (200 MHz) FT spectrometers in the indicated
solvent. Chemical shifts are expressed in d units (ppm)
from tetramethylsilane. Column chromatographies were
packed with alumina Merck (70±230 mesh) and silica gel
Merck (70±230 mesh). Aluminum oxide TLC cards Fluka
(aluminum oxide precoated aluminum cards with ¯uor-
escent indicator at 254 nM) and silica gel TLC cards
Fluka (silica gel precoated aluminum cards with ¯uor-
escent indicator at 254 nM) were used for thin-layer chro-
matography (TLC). Developed plates were visualized by
spectroline ENF 260C/F UV apparatus. Organic solu-
tions were dried over anhydrous sodium sulfate. Con-
centration and evaporation of the solvent after reaction or
extraction was carried out on a rotary evaporator Buchi
Rotavapor operating at reduced pressure. Elemental ana-
lyses were performed by laboratories of Dr. M. Zancato,
Dipartimento di Scienze Farmaceutiche, University of
Padova (Italy). Analytical results were within Æ0.4% of
the theoretical values. Ethyl 1H-pyrrole-2-carboxylate10
and 4-¯uoro-3-nitrobenzenesulfonyl chloride11 were pre-
pared according to the literature.
1
(CO), 3300 and 3440 cm (NH2). Anal. C13H13N3O6S
(339.32), C, H, N, S.
Ethyl 1-[(3,4-diaminophenyl)sulfonyl]-1H-pyrrole-2-car-
boxylate (10). A stirred solution of 9 (1.70 g, 0.005 mol)
in glacial acetic acid (21 mL) was treated portionwise with
iron powder (1.5g) while heating at 60ꢀC under stirring.
Reaction mixture was maintained at 60ꢀC for 2 h, then
evaporated to dryness. The residue was mixed with ice
water and extracted with ethyl acetate. The organic layers
were collected, washed with brine and dried. Removal of
the solvent gave a residue which was puri®ed by column
chromatography (silica gel/ethyl acetate) to yield 10
(1.0 g, 65%), mp 155±156 ꢀC (toluene). 1H NMR
(DMSO-d6): d 1.20 (t, J=7.0 Hz, 3H, COO CH2CH3),
4.16 (q, J=7.0 Hz, 2H, COOCH2CH3), 4.97 (broad s,
2H, NH2PhNH2, disappeared on treatment with D2O),
5.65 (broad s, 2H, NH2PhNH2, disappeared on treat-
ment with D2O), 6.34 (3 line m, 1H, H4-pyrrole), 6.57
(d, J=8.4 Hz, 1H, H5-benzene), 6.98 (dd, J=1.5 and
3.6 Hz, 1H, H3-pyrrole), 7.02 (d, J=2.2 Hz, 1H, H2-
benzene), 7.10 (dd, J=2.0 and 8.4 Hz, 1H, H6-benzene),
Ethyl 1-[(4-¯uoro-3-nitrophenyl)sulfonyl]-1H-pyrrole-2-
carboxylate (8). To a stirred mixture of potassium tert-
butoxide (1.34 g, 0.012 mol) and 18-crown-6 (0.28 g,
0.0011 mol) in anhydrous THF (21 mL) was added
dropwise a solution of ethyl 1H-pyrrole-2-carboxylate
(1.39 g, 0.010 mol) in the same solvent (21 mL). Stirring
was maintained for 15 min. The suspension of potassium
salt was cooled to 0 ꢀC and added by small portions to a
solution of 4-¯uoro-3-nitrobenzenesulfonyl chloride
(2.57 g, 0.010 mol) in anhydrous THF (21 mL). Reaction
was stirred at room temperature for 3.5 h, then con-
centrated to a small volume and extracted with ethyl
acetate. Organic extracts were washed with brine and
dried. Removal of the solvent gave a residue which
was puri®ed on silica gel column chromatography (di-
chloromethane). First fractions gave traces of 4-¯uoro-
3-nitrobenzenesulfonyl chloride. Second fractions a€orded
7.61 ppm (m, 1H, H5-pyrrole). IR (Nujol): n 1705 (CO),
1
3270, 3350, 3420 and 3450 cm
C13H15N3O4S (309.34), C, H, N, S.
(NH2). Anal.
Ethyl 1-[(1H-benzimidazol-2(3H)one-5-yl)sulfonyl]-1H-
pyrrole-2-carboxylate (5). Bis(trichloromethyl) carbon-
ate (triphosgene) (0.15 g, 0.0005 mol) in dichloromethane
(10 mL) was dropped into a solution of 10 (0.31 g,
0.0010mol) and triethylamine (0.12 g, 0.0012mol) in the
same solvent (30 mL). Reaction was stirred at room
temperature for 24h, then diluted with water and
extracted with ethyl acetate. The organic layer was
washed with brine, dried and evaporated to dryness. The
1
8 (2.2 g, 65%); thick oil. H NMR (CDCl3): d 1.29 (t,
J=7.1 Hz, 3H, COO CH2CH3), 4.21 (q, J=7.1 Hz, 2H,
COOCH2CH3), 6.38 (3 line m, 1H, H4-pyrrole), 7.08
(dd, J=1.4 and 3.6 Hz, 1H, H3-pyrrole), 7.49 (t,
J=9.3 Hz, 1H, H5-benzene), 7.71 (unresolved dd, 1H,
H5-pyrrole), 8.32±8.43 (8 line m, 1H, H6-benzene),
8.69 ppm (dd, J=2.5 and 6.8 Hz, 1H, H2-benzene). IR
(Nujol): n 1710cm 1 (CO). Anal. C13H11FN2O6S (342.29),

R. Silvestri et al. / Bioorg. Med. Chem. 8 (2000) 2305±2309
2309
crude residue was puri®ed on silica gel column (ethyl
acetate) to yield 5 (0.31 g, 92%); mp >237±238ꢀC (eth-
anol). H NMR (DMSO-d6): d 1.19 (t, J=7.0Hz, 3H,
Molecular modeling
1
Molecular modeling studies were performed using the
SYBYL13 software package running on a Silicon Graphics
R10000 workstation. Intramolecular and intermolecular
energies were calculated using the molecular mechanics
Tripos force ®eld14 including the electrostatic contribution.
Atom centered partial charges were calculated accord-
ing to the Gasteiger±Huckel method.15,16 Geometry
optimizations were realized with the SYBYL/MAX-
IMIN2 minimizer by applying the BFGS algorithm17
and setting a root-mean-square gradient of the forces
acting on each atom at 0.05 kcal/mol A as a con-
vergence criterion. Model building and docking of 3
into the RT binding site of non-nucleoside inhibitors
(extracted from the RT/R90385 complex solved at 2.4 A
resolution8) are detailed in a previous paper.9 Complexes
of compounds 5±7 with RT were constructed starting
from the docked conformation of 3 and submitted to
geometry-optimization.
COO CH2CH3), 4.15 (q, J=7.0 Hz, 2H, COOCH2CH3),
6.43 (3 line m, 1H, H4-pyrrole), 7.04 (dd, J=2.0 and
3.4 Hz, 1H, H3-pyrrole), 7.14 (d, J=8.3 Hz, 1H, H5-
benzene), 7.52 (u, 1H, H2-benzene), 7.64 (dd, J=1.9 and
8.3 Hz, 1H, H6-benzene), 7.82 (dd, J=2.0 and 3.1 Hz,
1H, H5-pyrrole), 11.11 and 11.31 ppm (two broad s, 2H,
NHCONH, disappeared on treatment with D2O). 13C
NMR (DMSO-d6): d 14.00 (COOCH2CH3), 60.52 (COO
CH2CH3), 107.91, 108.37, 110.73, 121.83, 122.68, 124.43,
129.04, 129.29 and 134.62 (aromatic C), 155.23 and 158.28
1
(COOEt and NHCONH). FT-IR: n 1713 and 1731cm
(CO). Anal. C14H13N3O5S (335.33), C, H, N, S.
Ethyl 1-[(1H-benzimidazol-5(6)-yl)sulfonyl]-1H-pyrrole-
2-carboxylate (6). A mixture of 10 (1.00 g, 0.0032 mol)
and formic acid (98±100%, 0.21 mL) was heated at
100 ꢀC for 2 h. After cooling the mixture was made alka-
line with 10% aqueous NaOH and shaken with ethyl ace-
tate. The organic layer was washed with brine and dried.
Removal of the solvent furnished the crude product which
was passed through a silica gel column (ethyl acetate) to
Acknowledgements
yield 6 (1.0g, 98%); mp 139±140 ꢀC (toluene). H NMR
The authors thank Italian Ministero della Sanita- Isti-
tuto Superiore di Sanita- X Progetto AIDS 1997 (grants
Nos 40A.0.06 and 40A.0.55), Italian MURST (40%
funds) and Regione Autonoma Sardegna (Progetto
Biotecnologie) for ®nancial support.
1
(DMSO-d6): d 1.17 (t, J=7.1Hz, 3H, COOCH2CH3),
4.14 (q, J=7.1 Hz, 2H, COOCH2CH3), 6.45 (m, 1H), 7.06
(m, 1H), 7.82 (m, 2H), 7.90 (m, 1H), 8.36 (broad, 1H),
8.55 (broad, 1H), 13.07 (broad s, 1H, NH, disappeared
1
on treatment with D2O). IR (Nujol): n 1720 cm (CO).
Anal. C14H13N3O4S (319.33), C, H, N, S.
References
Ethyl 1-[(1H-benzotriazol-5(6)-yl)sulfonyl]-1H-pyrrole-2-
carboxylate (7). An aqueous solution of sodium nitrite
(0.45 g, 0.0066 mol, 1.5 mL) was added in one portion to
a water cooled solution of 10 (1.86 g, 0.0060 mol) in
50% aqueous acetic acid (100 mL). The reaction was
stirred for 1 h, then mixture was poured on crushed ice
while stirring for some minutes. The solid which formed
was ®ltered, washed with water and dried. Crude pro-
duct was passed through a silica gel column (ethyl/ace-
tate) to a€ord 7 (1.5 g, 78%); mp 130±131ꢀC (toluene/
cyclohexane). 1H NMR (CDCl3): d 1.22 (t, J=7.1 Hz, 3H,
COOCH2CH3), 4.15 (q, J=7.1 Hz, 2H, COOCH2CH3),
6.38 (3 line m, 1H, H4-pyrrole), 7.10 (dd, J=1.8 and
3.7Hz, 1H, H3-pyrrole), 7.79 (dd, J=1.8 and 3.1Hz, 1H,
H5-pyrrole), 7.96 (m, 2H, benzene), 8.77 ppm (m, 1H,
benzene). IR (Nujol): n 1700 (CO), 3320 cm 1 (NH). Anal.
C13H12N4O4S (320.32), C, H, N, S.
1. De Clercq, E. AIDS Res. Hum. Retrovirus 1992, 8, 119.
2. Young, S. D. Perspect. Drug Discovery Des 1993, 1, 181.
3. Mitsuya, H.; Weinhold, K. J.; Furman, P. A.; St. Clair, M.
H.; Nusino€-Lehrman, S.; Gallo, R. C.; Bolognesi, D.; Barry,
D. W.; Broder, S. Proc. Natl. Acad. Sci. USA 1985, 82, 7096.
4. Merluzzi, V. J.; Hargrave, K. D.; Labadia, M.; Grozinger,
K.; Skoog, M.; Wu, J. C.; Shih, C.-K.; Eckner, R. J.; Koup, R.
A.; Sullivan, J. L. Science 1990, 250, 1411.
5. De Clercq, E. Clin. Microbiol. Rev. 1995, 8, 200.
6. Kohlstaedt, L. A.; Wang, J.; Friedman, J. M.; Rice, P. A.;
Steitz, T. A. Science 1992, 256, 1783.
7. Artico, M.; Silvestri, R.; Massa, S.; Loi, A. G.; Corrias, S.;
Piras, G.; La Colla, P. J. Med. Chem. 1996, 39, 522.
8. Ren, J.; Esnouf, R.; Garman, E.; Somers, D.; Ross, C.;
Kirby, I.; Keeling, J.; Darby, G.; Jones, Y.; Stuart, D.; Stam-
mers, D. Nat. Struct. Biol. 1995, 2, 293.
9. Artico, M.; Silvestri, R.; Pagnozzi, E.; Bruno, B.; Novel-
lino, E.; Greco, G.; Massa, S.; Ettorre, A.; Loi, A. G.; Scintu,
F.; La Colla, P. J. Med. Chem. 2000, 43, 1886.
10. Bailey, D. M.; Johnson, R. E.; Albertson, N. F. Org.
Synth. 1971, 51, 100.
Biological evaluations
Activity of the compounds against HIV-1 multiplication
in acutely infected cells was based on the inhibition of
virus-induced cytopathicity in MT-4 and C8166 cells,
respectively. According to a previously reported pro-
cedure,7 the number of viable cells was determined by the
3-(4,5 - dimethylthiazol - 2 - yl) - 2,5 - diphenyltetrazolium
bromide (MTT) method. Cytotoxicity of the compounds
was evaluated in parallel with their antiviral activity. It
was based on the viability of mock-infected cells, as
monitored by the MTT method. Anti-RT assays were
performed as described in a previous paper.7
11. Dehn, J. W., Jr.; Gancher, E.; Resniek, P.; Kuhenfuss,
H.J. Patent Fr. 1,412,060 (1965). Chem. Abstr. 1996, 65,
7326d.
12. Cotarca, L.; Delogu, P.; Nardelli, A.; Sunjic, V. Synthesis,
1996, 553.
13. SYBYL Molecular Modeling System (version 6.2), Tripos
Assoc. St. Louis, Missouri, USA.
14. Clark, M.; Cramer, R. D., III; Van Opdenbosch, N. J.
Comput. Chem. 1989, 10, 982.
15. Gasteiger, J.; Marsili, M. Tetrahedron 1980, 36, 3219.
16. Purcel, V. P.; Singer, J. A. J. Chem. Eng. Data 1967, 12, 235.
17. Head, J.; Zerner, M. C. Chem. Phys. Lett. 1985, 122, 264.