Structure activity of 3-Aryl-1,3-diketo-containing compounds as HIV-1 integrase inhibitors

Article abstract of DOI:10.1021/jm020037p

The 4-aryl-2-hydroxy-4-oxo-2-butenoic acids and their isosteric tetrazoles are among an emerging class of aryl β-diketo (ADK)-based agents which exhibit potent inhibition of HIV-1 integrase (IN)-catalyzed strand transfer (ST) processes, while having much

Full text of DOI:10.1021/jm020037p

3184
J . Med. Chem. 2002, 45, 3184-3194
Str u ctu r e Activity of 3-Ar yl-1,3-d ik eto-Con ta in in g Com p ou n d s a s HIV-1
In tegr a se In h ibitor s1
Godwin C. G. Pais,† Xuechun Zhang,† Christophe Marchand,‡ Nouri Neamati,‡ Kiriana Cowansage,‡
Evguenia S. Svarovskaia,§ Vinay K. Pathak,§ Yun Tang,† Marc Nicklaus,† Yves Pommier,‡ and
Terrence R. Burke, J r.*,†
Laboratory of Medicinal Chemistry, Laboratory of Molecular Pharmacology, and HIV Drug Resistance Program, Center for
Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, Maryland 21702-1201
Received J anuary 24, 2002
The 4-aryl-2-hydroxy-4-oxo-2-butenoic acids and their isosteric tetrazoles are among an
emerging class of aryl â-diketo (ADK)-based agents which exhibit potent inhibition of HIV-1
integrase (IN)-catalyzed strand transfer (ST) processes, while having much reduced potencies
against 3′-processing (3′-P) reactions. In the current study, L-708,906 (10e) and 5CITEP (13b),
which are two examples of ADK inhibitors that have been reported by Merck and Shionogi
pharmaceutical companies, served as model ADK leads. Structural variations to both the “left”
and “right” sides of these molecules were made in order to examine effects on HIV-1 integrase
inhibitory potencies. It was found that a variety of groups could be introduced onto the left
side aryl ring with maintenance of good ST inhibitory potency. However, introduction of
carboxylic acid-containing substituents onto the left side aryl ring enhanced 3′-P inhibitory
potency and reduced selectivity toward ST reactions. Although both L-708,906 and 5CITEP
show potent inhibition of IN in biochemical assays, there is a disparity of antiviral activity in
cellular assays using HIV-1-infected cells. Neither 5CITEP nor any other of the indolyl-
containing inhibitors exhibit significant antiviral effects in cellular systems. Alternatively,
consistent with literature reports, L-708,906 does provide antiviral protection at low micromolar
concentrations. Interestingly, several analogues of L-708,906 with varied substituents on the
left side aryl ring, while having good inhibitory potencies against IN in extracellular assays,
are not antiviral in whole-cell systems.
In tr od u ction
Human immunodeficiency virus type 1 (HIV-1) en-
codes three enzymes which are required for viral
replication: reverse transcriptase, protease, and inte-
grase (IN). Although drugs targeting reverse tran-
scriptase and protease are in wide use and have shown
effectiveness particularly when employed in combina-
tion,2 toxicity and development of resistant strains have
limited their usefulness.3 Impetus, therefore, exists for
the discovery of new agents directed against alternate
sites in the viral life cycle. IN has emerged as an
attractive target, because it is necessary for stable
infection4 and homologous enzymes are lacking in the
human host. The function of IN is to catalyze integration
of proviral DNA, resulting from the reverse transcrip-
tion of viral RNA, into the host genome. This is achieved
in a stepwise fashion by endonucleolytic processing of
proviral DNA within a cytoplasmic preintegration com-
plex (termed 3′-processing or “3′-P”), followed by trans-
location of the complex into the nuclear compartment
where integration of 3′-processed proviral DNA into host
DNA occurs in a “strand transfer” (ST) reaction. Al-
though numerous agents potently inhibit 3′-P and ST
F igu r e 1. General aryl diketo-based inhibitor (1) with two
recently reported examples.
in extracellular assays that employ recombinant IN and
viral long terminal repeat oligonucleotide sequences,5
often such inhibitors lack inhibitory potency when
assayed using fully assembled preintegration com-
plexes6 or fail to show antiviral effects against HIV-
infected cells.7 Recently, however, a class of IN inhibi-
tors has emerged, typified by an aryl â-diketo motif
(ADK (1), Figure 1). Variations of this structural theme
have been utilized by Shionogi8,9 and Merck10-12 phar-
maceutical companies to prepare inhibitors (for ex-
ample, 5CITEP13 and L-708,906,14 Figure 1), some of
which potently block integration in extracellular assays
and exhibit good antiviral effects against HIV-infected
cells. Further studies have indicated that, unlike nu-
merous other IN inhibitors whose antiviral effects can
be attributable to non-integrase-dependent phenom-
ena,15,16 members of the ADK family disrupt viral
* To whom correspondence should be addressed: Laboratory of
Medicinal Chemistry, Center for Cancer Research, NCI-Frederick, P.O.
Box B, Bldg. 376 Boyles St., Frederick, MD 21702-1201. Tel: (301)
846-5906. Fax: (301) 846-6033. E-mail: tburke@helix.nih.gov.
† Laboratory of Medicinal Chemistry.
‡ Laboratory of Molecular Pharmacology.
§ HIV Drug Resistance Program.
10.1021/jm020037p This article not subject to U.S. Copyright. Published 2002 by the American Chemical Society
Published on Web 06/06/2002

Activity of 3-Aryl-1,3-diketo-Containing Compounds
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15 3185
Sch em e 1a
a
(i) (CO2Et)2, NaH, toluene (60 °C, 3 h)(for 2e, 110 °C, 1 h); (ii) 1 N NaOH, dioxane (rt, 1 h).
Sch em e 2a
a
(i) (CO2Et)2, NaOEt, THF (rt, 3 h; 50 °C, 18 h); (ii) KOH, EtOH (reflux 15 h) or 1 N NaOH, THF-EtOH (1:1) (rt 1 h) for 6c.
Sch em e 3a
infectivity in a manner consistent with inhibition of
integration.14,16,17 Finally, evidence has been presented
that at least some ADK-based inhibitors show selective
inhibition of the ST reaction relative to the 3′-P step
and may function by competing with host DNA in
binding to the IN‚proviral DNA complex.14,18,19 Although
ADK-motifs currently present some of the most promis-
ing starting points for inhibitor development,20-22 few
stucture-activity relationship (SAR) studies have been
reported on this genre of compounds.23 Accordingly,
herein is reported an examination of the structural
features of the ADK family as they relate to IN inhibi-
tory potency and selectivity.
a
(i) (CO2Et)2, NaOEt, THF (rt, 3 h; 50 °C, 18 h) or NaH, toluene
(for 9d , 110 °C, 1 h); (ii) KOH, EtOH (reflux 15 h) or 1 N NaOH,
THF-EtOH (1:1) (rt, 1 h) for 10c,d .
Syn th esis
milder conditions were employed (1 N NaOH in THF-
EtOH at room temperature).23 Heteroaryl and ferrocene-
containing acids 10a -d and 3,5-dibenzyloxy derivative
10e were prepared in a similar fashion (Scheme 3).
4-Ar yl-3-h yd r oxy-4-oxo-3-(1H-tetr a zol-5-yl)-p r o-
p en on e Der iva t ives (12, 13, 15, a n d 16). Acetyl
indoles 8a ,b,f,g were coupled with 1126 in the presence
of LHMDS to provide p-methoxybenzyl-protected tet-
razoles 12a ,b,f,g. The final products 13a ,b,f,h were
obtained by the removal of the tetrazole protection using
TFA with ethanedithiol as a scavenger (Scheme 4). For
13h , hydrogenolytic cleavage of the benzyl ester was
accomplished prior to treatment with TFA. Tetrazole
14 was synthesized from 3,5-dimethoxybenzyl bromide
in a fashion similar to the synthesis of 11. Treatment
of 8e with 11 gave the protected tetrazole derivative
16b. Attempts to selectively remove the p-methoxyben-
zyl group of 16b were not successful (Scheme 5).
Eva lu a tion of An tivir a l Activity. A single-cycle
replication assay was utilized to measure the antiviral
activity of selected compounds, wherein an envelope-
deficient HIV-1-based retroviral vector containing the
4-Ar yl-2-h yd r oxy-4-oxo-2-bu ten oic Acid s. Synthe-
sis of 4-aryl-4-oxo-2-hydroxy-2-butenoic acids occurred
with the oxalylation of the corresponding aryl ketones
in the presence of base, followed by either alkaline or
acidic hydrolysis (Schemes 1-3).23,24 For analogues
containing a phenyl aryl portion and a single 2,4-
dioxobutanoyl side chain (4a -e), synthesis according to
the literature procedures provided aryl ketones (2a -
e). These were coupled with diethyl oxalate in the
presence of NaH to give intermediate ethyl esters (3a -
e) which were converted to free acids (Scheme 1). Among
the methods investigated for ethyl ester hydrolysis,
aqueous 1 N NaOH in dioxane proved to be the most
effective.
Synthesis of bis-[4-(2-hydroxy)-4-oxo-2-butenoic acid]-
containing compounds 7a -c was achieved in a similar
fashion by coupling bis-(methyl ketones) 5a -c with
diethyl oxalate in the presence of NaOEt,25 with hy-
drolysis of the resulting bis-(ethyl esters) 6a -c being
subsequently carried out in the presence of KOH in
refluxing EtOH (Scheme 2). Note that for product 7c,

3186 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15
Pais et al.
Sch em e 4a
the 3′-processing reaction). The distance between the
two Mg2+ ions remained constant throughout the mo-
lecular dynamics simulation at approximately 7 Å.27
Resu lts a n d Discu ssion
Our interest in the ADK class of IN inhibitors began
with the disclosure of the X-ray crystal structure of
5CITEP (13b) complexed to the catalytic D64, D116, and
E152 residues of the IN core domain.13 This provided
preliminary evidence that ADK-containing agents may
potentially afford promising leads for the development
of IN inhibitors.22 Such expectations were supported by
subsequent reports that structurally related ADK-
containing compounds exhibit potent IN inhibition and
elicit good antiviral effects in HIV-infected cells.14,18
Although both Shionogi’s 5CITEP and Merck’s ADK-
based inhibitors share a common â-diketopropyl central
chain, they differ in their “left” and “right” portions
(Figure 1). Since only limited structure-activity or
synthetic studies of these inhibitors have been re-
ported,23 the current study was undertaken to examine
aspects of both the left and right portions of the
molecules.
In h ibitor y P oten cies in HIV-1 In tegr a se As-
sa ys: Mod ifica tion s to th e Left Sid e of 5CITEP . In
the report of 5CITEP (13b) complexed with the IN
catalytic domain, inhibitory data against IN were
minimal and did not include antiviral potency in HIV-
infected cells.13 Therefore, this previously disclosed
inhibitor was prepared in order to determine its inhibi-
tory profile against 3′-P and ST as well as its antiviral
potency in HIV-infected cells. Additionally, the impor-
tance of the chloro substituent at the indolyl 5-position
was examined through the preparation of agents (13a ,
13f, and 13h ) whose functionalities varied at the
5-position. Inhibitory values measured in an extracel-
lular integrase assay are shown in Table 1. Antiviral
potencies in HIV-infected cells were also determined
(Table 2) for analogues exhibiting IC50 e 10 µM in this
assay. We found that 5CITEP (13b) exhibits potent
inhibition of ST, while it is much less effective for
inhibiting 3′-P (approximately 50-fold). Substitution of
the 5-Cl with hydrogen (13a ) decreases 3′-P inhibitory
potency, whereas fluorine (13f) exhibits 3′-P inhibitory
potency equivalent to that of the parent chlorine.
Substitution with either hydrogen or fluorine has little
effect on ST inhibitory potency. Replacement of the 5-Cl
with a carboxyl group (13h ) significantly decreases the
a
(i) LHMDS, THF (-20 °C, 2 h, -78 °C to rt, 1 h, rt, 2 h); (ii)
TFA, ethane dithiol-H2O (1:1) (rt, 20 h) [for 12g, H2/Pd‚C, THF
then (ii)]. Note: For 13g, X ) HO2C.
firefly luciferase reporter gene (pNluc) was cotransfected
into 293T cells with vesicular stomatitis virus G glyco-
protein (VSV-G) expressing plasmid (CMV-VSV-G).
Virus harvested from transfected cells was used to infect
target cells in the presence or absence of the compounds
tested. The ability of compounds to inhibit viral replica-
tion was measured by determining the amount of
luciferase activity in the infected cells. Additionally,
concentrations of compounds that resulted in 50%
inhibition of HIV-1 infection (IC50 values) were deter-
mined for compounds that displayed antiviral activity
at a concentration of 25 µM (Table 2). For these latter
determinations, single-cycle HIV-1 replication assays
were performed in the presence of a series of inhibitor
concentrations (0.05, 0.5, 5, and 50 µM). To determine
a relative inhibition of HIV-1 infection at each concen-
tration, luciferase activity of each sample was normal-
ized to the untreated control.
Molecu la r Mod elin g. A model of the IN core com-
plexed with viral DNA was initially constructed utilizing
all available experimental evidence and refined by
molecular dynamics simulation.27 This hypothetical
model was intended to represent IN immediately fol-
lowing the first catalytic reaction (3′-processing). The
model was refined by placing two Mg2+ ions into the
active site, which is a cleft formed partly by both the
protein and DNA. The location of these metal ions was
based on experimental evidence and the hypothesis that
a second metal ion was likely to be carried into the
HIV-1 IN active site by viral DNA binding. 28 One Mg2+
ion coordinated with the carboxyl groups of Asp64 and
Asp116. The other metal ion was bound to the carboxyl
group of Glu152, the carbonyl group of Asn155, and the
phosphate group of the last adenosine of the viral DNA
(proximal to the G-T nucleotides which are cut during
Sch em e 5a
a
(i) LHMDS, THF -20 °C, 2 h, -78 °C to rt, 1 h; rt 2 h.

Activity of 3-Aryl-1,3-diketo-Containing Compounds
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15 3187
Ta ble 1. Inhibitory Potencies As Measured in an Extracellular HIV-1 Integrase Assaya
a
b
Values were determined as described in the Experimental Section. Structures as described in Scheme 2.
Ta ble 2. Antiviral Data for Selected Inhibitors As Measured in a Single-Cycle Replication Assay Using Envelope-Deficient HIV-1a
% control % control
experiment 2 experiment 2
no.
experiment 1
comment
no.
experiment 1
comment
cytotoxicb
4a
4b
4d
7a
7b
7c
9b
92
14
261
98
133
61
104
103
25
208
95
128
94
63
10a
10b
10d
10e
13a
13b
13f
1
91
36
0
85
69
19
87
69
70
IC50 ) 0.6 µM
cytotoxicb
IC50 ) 2.3 µM
7
116
125
65
a
Compounds exhibiting ST IC50 values of e10 µM in extracellular HIV integrase assays (Table 1) were selected for evaluation and
tested initially at a concentration of 25 µM as described in the Experimental Section. Compounds 4b and 10e, which achieved g50%
b
inhibition at 25 µM, were examined over a concentration range to determine the IC50 values. Cytotoxicity was determined by visual
inspection of cells.
inhibitory potency against ST while approximately
doubling potency against 3′-P. This resulted in a com-
plete loss of ST/3′-P selectivity.
Mod ifica tion s to th e Righ t Sid e. L-708,906 (10e)
and 5CITEP (13b), while sharing a common 1,3-dike-
topropyl moiety, differ both in their left aryl portions

3188 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15
Pais et al.
and in their right side functionality. The former pos-
sesses a carboxylic acid group, and the latter possesses
a tetrazole group. It is known that tetrazoles can serve
as isosteric replacements for carboxyl groups.29,30 How-
ever, the biological effects of such an interchange are
complicated in the ADK class of IN inhibitors due to
the potential importance of tetrazole nitrogens for the
binding of 5CITEP to the enzyme as indicated by X-ray
crystallography.13 Therefore, replacement of the tetra-
zole group with carboxyl functionality was examined for
both 5CITEP (13b) and its des-chloro congener (13a ).
The resulting analogues (10b and 10a , respectively)
exhibited little change in either 3′-P or ST inhibition
potencies (Table 1). The blockade of the carboxyl hy-
droxyl of the 5CITEP analogue 10b by conversion to its
ethyl ester (9b) was accompanied by maintenance of the
significant ST inhibitory potency and selectivity over 3′-
P. This indicates that an acidic proton may not be
required for ST inhibitory potency. The fact that a
blockade of acidic protons of tetrazole-containing ana-
logues (13a ,b), with either 4-methoxybenzyl groups
(12a ,b) or a 3,5-dimethoxybenzyl group (15a ), resulted
in a significant loss of ST inhibitory potency may be due
to steric hindrance rather than a loss of acidic protons.
phenyl ring of 4e with a 4-quinolyl ring (10c) resulted
in a dramatic loss of inhibitory potency. This is in
contrast to the good ST inhibitory potency shown by the
indolyl analogue 10a .
Although above outlined modifications to L-708,906
(10e) affected the potency of ST inhibition, the selectivi-
ties of ST versus 3′-P were maintained. In a further set
of analogues, additional 2,4-diketobutyric acid groups
were added at the 3- or 4-position of the left side aryl
ring (7a or 7b, respectively). Interestingly, while ST
inhibitory potency was largely unaffected, these bifunc-
tional analogues displayed enhanced 3′-P inhibitory
potencies, such that selectivity toward ST was very
much reduced. Adding a second diketo acid side chain
to quinolyl-containing 10c, which was essentially inac-
tive, gave 7c which displayed greatly restored inhibitory
potency. The loss of ST to 3′-P selectivity by these
bifunctional analogues differentiates them from other
ADK inhibitors which are generally characterized by
high ST selectivity. Potential mechanistic considerations
of bis-diketo acid inhibitors are discussed elsewhere.19
An tivir a l Effects. While potent inhibition in extra-
cellular IN assays has been reported for an extensive
array of compounds, demonstration of antiviral effects
against HIV-infected cells for agents that exhibit good
potency in these extracellular enzyme assays has been
more elusive. The ability of certain ADK-based IN
inhibitors to elicit potent antiviral effects by mecha-
nisms consistent with intracellular inhibition of IN has
set this class apart.14,16,17
An a logu es of L-708,906. A second class of ADK-
containinginhibitors,as exemplified byMerck’s L-708,90614
(Figure 1, designated as 10e in Table 1), is characterized
by a carboxyl group on the right side and multiple aryl
rings on the left side. The biological properties of this
class have been more extensively detailed than those
of 5CITEP. It has been shown that L-708,906 potently
inhibits HIV-1 IN-catalyzed ST processes while having
little effect on 3′-P catalysis.18 Inhibition of viral replica-
tion in HIV-1-infected cells by L-708,906 is achieved at
low micromolar concentrations, and resistant viral
strains bear mutations which map to integrase. These
mutations include M154I, S153Y, and T66I which are
proximal to active site Glu152 and Asp64 residues,
respectively.18
A total of 12 compounds were tested for their ability
to inhibit HIV-1 infection. The compounds were prese-
lected for antiviral evaluation based on the ST inhibitory
potencies (IC50 values e 10 µM) as listed in Table 1. It
should be noted that Mn2+ rather than Mg2+ was
utilized for in vitro assays, because the soluble mutant
integrase enzyme employed functions more efficiently
with this metal ion than with Mg2+. Although in vivo
the cation used by integrase is unclear, it may be Mg2+
,
and for this reason, IC50 values obtained using Mn2+
may not accurately reflect in vivo inhibition. Initially,
the compounds were tested at 25 µM concentrations
with the results being summarized in Table 2. L-708,906
(10e) was used as a control because of its known ability
to inhibit HIV-1 integration in vitro and in vivo.17,18 In
the current assay, 10e at 25 µM reduced HIV-1 infection
to an average of 13% of the untreated control, indicating
that this compound was active in inhibiting HIV-1
replication. For compounds exhibiting antiviral activity
at 25 µM concentrations (4b and 10e), further experi-
ments were conducted to determine antiviral IC50
values. These experiments indicated that IC50 values
for 4b and 10e were 0.6 and 2.3 µM, respectively. The
IC50 value for 10e (2.3 µM) was very similar to the
previously reported IC50 value (2.5 µM).18 Interestingly,
although 4b is structurally related to 10e, it exhibits a
lower IC50 value (0.6 µM) suggesting that it may be more
effective at inhibiting HIV-1 replication than 10e. Two
compounds (10a and 10d ) displayed deleterious effects
on cell viability as determined by visual examination
of cell cultures following incubation with these com-
pounds during infection. For these agents, reduction in
luciferase activity was most likely due to cytotoxicity
(Table 2).
Further studies, using related analogues bearing
pyrrole-based left side groups, provide evidence that this
class of ADK inhibitor may function by competing for
host DNA binding to the IN‚viral DNA complex.14
Finally, a structure-activity investigation based on a
pyrrole-containing lead ADK inhibitor has presented a
focused examination of specific aspects of this family.23
Our current study is complementary to this latter report
in exploring a wider range of structural diversity in the
left side portion of the ADK theme.
Consistent with literature findings,18 L-708,906 ex-
hibited potent inhibition of ST with no effect on 3′-P up
to 1000 µM (10e, Table 1). Removal of one benzyloxy
group (compound 4b) resulted in a slight increase in ST
inhibitory potency and some loss of selectivity relative
to 3′-P. Removal of the remaining 3-benzyloxy group (4e)
gave a greater than 50-fold loss of ST inhibitory potency,
while introduction of a 3-bromo substituent (4a ) gave a
significant restoration of potency. The importance of
functionality at the aryl 3-position was shown further
when the replacement of the benzyloxy group in 4b with
a benzoylamino group (4c) reduced ST inhibitory po-
tency by approximately 100-fold. Aryloxy substituents
at other sites, such as the 4-position (4d ), also reduced
ST inhibitory potency. Replacement of the unsubstituted

Activity of 3-Aryl-1,3-diketo-Containing Compounds
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15 3189
F igu r e 2. (A) Docking model of L-708,906 in the active site of an IN core‚viral DNA complex, prepared by FlexX and refined
with MD simulation (500 ps). (B) Docking model of 5CITEP at the active site of an IN core‚viral DNA complex, derived by FlexX
and refined with MD simulation (300 ps). (C) Comparison of FlexX-derived docking model with the X-ray-determined 5CITEP
bound to the free IN core domain (PDB entry: 1QS4). These figures were produced with InsightII 2000.40 All balls stand for Mg2+
ions. Residues from an IN‚DNA complex are colored in light blue; residues from a free dynamics-averaged IN core domain dimer
are colored in purple, and residues from the X-ray structure 1QS4 are shown in yellow. The last two nucleotides C-A of viral
DNA are shown in orange.
Molecu la r Mod elin g. By utilizing the hypothetical
model of IN complexed with viral DNA (developed as
described above), FlexX-derived docking models were
generated for the binding of both L-708,906 (10e) and
5CITEP (13b) within the active site. Results of these
FlexX-derived docking models were quite similar for
L-708,906 and 5CITEP (Figure 2). In both cases, coor-
dination of the second Mg2+ ion with the common
â-diketopropyl moiety differed only slightly. The car-
boxyl group of L-708,906 bound to the first Mg2+ ion in
addition to forming a hydrogen-bonding interaction with
Asp64 (Figure 2A). The central aromatic ring of
L-708,906 exhibited π-π stacking interactions with the
side chain of Tyr143, whereas the two benzyl rings
showed significant movement during the simulation.
Within the duration of the dynamics run, the tetrazole
group of 5CITEP failed to reach the Mg2+. Instead, it
formed hydrogen-bonding interactions with the phos-
phate group of the terminal adenosine of the viral DNA
(Figure 2B). The aromatic ring of 5CITEP formed edge-
to-face hydrophobic interactions with the aromatic plane
of Tyr143. This model of 5CITEP docked with the IN-
DNA complex exhibits geometry which is similar, but
not identical, to the published X-ray structure.13 In the
latter case, the â-diketopropyl functionality also points
toward the side chain of Glu152 (Figure 2C). However,
the X-ray complex was of IN before 3′-processing had
occurred. Following 3′-processing, one would expect that
the active site would undergo a change. From our
molecular dynamics simulations,27 it was found that the
side chains of residues Gln148, Glu152, Asn155, Lys156,
and Lys159 altered their conformations and became
more ordered after binding to viral DNA and the second
Mg2+ ion (Figure 2C).
Molecular modeling studies of potential binding to IN
complexed with viral DNA indicate that while the
carboxyl of L-708,906 can coordinate a second Mg2+ ion,
the tetrazole group of 5CITEP fails to reach this ion.
Instead, some interaction with the terminal phosphate
group of the viral DNA was observed during the
molecular dynamics simulation.
Con clu sion s
DKA-based IN inhibitors represent an emerging class
with potential relevance for the development of new
AIDS therapeutics. Work reported herein examines the
structural relationships of two important members of
this family, L-708,906 and 5ClTEP. Both compounds
share a common â-diketopropyl linker yet differ in both
their left aryl and right acidic portions. In regard to the
right side acidic functionality, it was observed that both
tetrazole and carboxyl moieties provide potent inhibition
in biochemical ST assays. Only carboxylic acid-bearing
agents 4b and 10e are antiviral within the concentra-
tion range tested. Reasons for the lack of antiviral
potency for tetrazole-containing inhibitors are not clear.
In regard to the left side functionality, significant
diversity is allowed for substituents on the phenyl
nucleus of L-708,906 and the indolyl nucleus of 5CITEP.
However, in the former case, substituents at the 3-posi-
tion are favored with the inhibitory potency of 3-mono-
benzyloxy analogue slightly exceeding that of parent 3,5-
dibenzyloxy L-708,906. Introduction of acidic functional-
ity into the left side aryl portion of compounds having
either carboxyl- or tetrazole-based right side acidic
functionality results in the enhancement of 3′-P inhibi-
tory potency and a reduction in 3′-P/ST selectivity.
As previously indicated, although both L-708,906 and
5CITEP show potent inhibition of IN in biochemical
assays, there is a disparity of antiviral activity in
cellular assays using HIV-1-infected cells. Neither
5CITEP nor any other of the indole-containing inhibi-
tors exhibit antiviral effects in cellular systems. Alter-
natively, L-708,906 does provide antiviral protection at
low micromolar concentrations. However, several ana-
logues of L-708,906 with varied substituents on the left
These modeling studies support similar binding modes
and mechanisms of action for L-708,906 and 5CITEP.
Both inhibitors form strong electrostatic interactions
within the active site which was modeled to represent
a state following 3′-processing but before strand trans-
fer. For both compounds, it seems reasonable to hypoth-
esize that they may inhibit strand transfer by occupying
the active site so as to hinder access of target DNA.

3190 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15
Pais et al.
necessary changes were made to have the Mg2+ ions and DNA
be considered as part of the putative active site.
side aryl ring while having good inhibitory potencies
against IN in biochemical assays, are not antiviral in
whole-cell systems. In summary, distinct activity dif-
ferences appear to exist between inhibitors related to
L-708,906 and those related to 5CITEP. Of the ana-
logues prepared in this study, those predicated on
L-708,906 appear more promising based on their anti-
viral activities.
Two of the primary docking models, one for each lead
compound, were selected to be refined further by MD simula-
tions. For these models, the programs CHARMM,36 version
c27b3, and the CHARMM all-atom force field, version 27,37,38
for protein and DNA were used on an SGI Origin 2000 and a
Beowulf-type Linux cluster in parallel.39 Force field parameters
for the ligands were developed with the software Quanta/
CHARMM 2000.40 All simulations were performed in explicit
aqueous solvent for 500 ps under the following conditions:
constant pressure (1 atm) and temperature (310 K), periodic
boundary conditions in a water box (sized 96 × 70 × 64 Å3),
12 Å cutoff for van der Waals interactions, and Particle Mesh
Ewald summation for electrostatic interactions. Docking mod-
els are shown in Figure 1.
Syn th esis. Gen er al Syn th etic Meth ods. Elemental analy-
ses were obtained from Atlantic Microlab Inc. (Norcross, GA),
and fast atom bombardment mass spectra (FABMS) were
acquired with a VG Analytical 7070E mass spectrometer under
the control of a VG 2035 data system. Where indicated, the
FABMS matrixes used were glycerol (Gly) or nitrobenzoic acid
(NBA). 1H-NMR data were obtained on a Brucker 250 MHz
or Varian 400 MHz spectrometer. The data are reported in
parts per million relative to TMS and referenced to the solvent
in which they were run. The solvent was removed by rotary
evaporation under reduced pressure, and silica gel chroma-
tography was performed using Merck silica gel 60 with a
particle size of 40-63 µm. Anhydrous solvents were obtained
commercially and used without further drying. HPLCs were
conducted using a Waters Prep LC4000 system having pho-
todiode array detection. Binary solvent systems were as
indicated where A ) 0.1% aqueous TFA and B ) 0.1% TFA in
acetonitrile. Either Vydac C18 Peptide and Protein (10 µm) or
Advantage C18 (5 µm) was used, in either preparative size (20
mm diameter × 250 mm long, with a flow rate of 10 mL/min)
or semipreparative size (10 mm diameter × 250 mm long, with
a flow rate of 2 mL/min).
Cou p lin g of Ar yl Meth yl Keton es. P r oced u r e A. To a
stirred solution of LHMDS (1.0 M in THF, 3.0 molar equiv) at
-78 °C was added dropwise aryl methyl ketone in anhydrous
THF, and the resulting mixture was stirred at -20 °C (2 h).
The reaction mixture was recooled to -78 °C, and a solution
of 11 in THF was added dropwise. The mixture was allowed
to warm to room temperature over 1 h and was stirred at room
temperature (2 h), and then the reaction was quenched by the
addition of saturated aqueous NH4Cl and subjected to an
extractive workup (EtOAc). Evaporation of the solvent pro-
vided a residue which was mixed with EtOAc to yield a bright
yellow solid. Recrystallization provided the desired product.
P r oced u r e B. To a stirred solution of NaOEt in anhydrous
THF at room temperature was added dropwise diethyl oxalate
in THF followed by aryl methyl ketone in THF. The resulting
orange-yellow mixture was stirred at room temperature (3 h),
and then at 50 °C (18 h). The solvent was removed under
reduced pressure to yield a yellow residue which was washed
with ether and collected by filtration. The collected solid was
washed with 1 N aqueous HCl and H2O, and then dried and
recrystallized to provide the desired final diketo product.
Exp er im en ta l Section
DNA Oligon u cleotid es. Oligonucleotides were purchased
from IDT Inc. (Coralville, IA) and purified on a 20% (19:1)
denaturing polyacrylamide gel using an UV shadow. Purified
oligonucleotides were 5′-end labeled by T4-polynucleotide
kinase (Gibco BRL/Life Technologies, Rockville, MD) as de-
scribed previously.31
HIV-1 In tegr a se In h ibition Assa y. The in vitro integra-
tion assay using recombinant HIV-1 integrase was performed
as described previously.31 Unless otherwise indicated, the
integrase-DNA complexes were preformed by mixing 400 nM
HIV-1 integrase with 10 nM 5′-end 32P-labeled 21-mer double-
stranded DNA template for 15 min on ice in a reaction buffer
containing 25 mM MOPS, pH 7.2, 25 mM NaCl, 7.5 mM
MnCl2, 0.1 mg/mL BSA, and 14.3 mM â-mercaptoethanol.
Inhibitors were then added to the reaction mixtures in a final
volume of 10 µL, and integration reactions were carried out
for 1 h at 37 °C. The reactions were quenched by adding 10
µL of denaturing loading dye. The samples were loaded onto
a 20% (19:1) denaturing polyacrylamide gel. The gels were
exposed overnight and analyzed using a Molecular Dynamics
Phosphorimager (Sunnyvale, CA).
Cells, Tr a n sfection , a n d In fection s. The 293T cells
(ATCC) were maintained in the presence of Dulbecco’s Modi-
fied Eagle’s Medium (Cellgro), 10% fetal calf serum (HyClone
Laboratories), penicillin (50 units/mL; Gibco), and streptomy-
cin (50 µg/mL; Gibco). The 293T cells were plated at a density
of 5 × 106 per 100 mm diameter dish and transfected using a
Transfection MBS Mammalian Transfection Kit (Stratagene).
Transfected cell supernatants were harvested 48 h after
transfection, clarified, and used to infect target cells. The 293T
target cells were plated at a density of 1 × 105 per 35 mm
diameter dish. The virus-containing medium was diluted 100-
fold and was used to infect the 293T target cells for 1 h as
previously described.28 The 293T target cells were incubated
with medium containing the test compounds for 4 h prior to
infection, 1 h during infection, and 24 h postinfection. Com-
pounds exhibiting ST IC50 values e10 µM in extracellular HIV
integrase assays (Table 1) were selected for evaluation. All
compounds were tested intially at 25 µM concentration, and
those agents which achieved g50% inhibition at 25 µM
(compounds 4b and 10e) were examined over a concentration
range to determine the IC50 values.
Lu cifer a se Assa y. Infected cells were washed with PBS
and lysed in 400 µL of reporter lysis buffer (Promega) 72 h
after infection. The samples were subjected to one freeze-thaw
cycle, and cell membranes were removed by centrifugation.
Luciferase activity was measured following the addition of 100
µL of substrate (Promega) to 20 µL of cell lysate using a TD20/
20 luminometer (Promega).
Molecu la r Mod elin g. The primary docking work was
performed on an SGI Octane workstation, running under the
IRIX64 version 6.5 operating system, using the molecular
modeling software package SYBYL 6.7.1.32 The IN core‚viral
DNA complex structure was extracted from the structural
model we had previously constructed of the complex with the
full-length IN with DNA33 and subjected to further refinement
by molecular dynamics (MD) simulation.27 The compounds
L-708,906 and 5ClTEP were generated in SYBYL and opti-
mized with ab initio calculations at the B3LYP/6-31G* level
of theory with the program Gaussian 98.34 L-708,906 and
5ClTEP were then docked into the active site of the IN core‚
viral DNA complex by means of flexible docking, using the
program FlexX35 (part of the SYBYL package), after the
P r oced u r e C. To aryl methyl ketone (1 molar equiv) in
toluene was added NaH (2.0 molar equiv), and the mixture
was stirred (10 min). Then diethyl oxalate (1.5 molar equiv)
was added dropwise, and the mixture was stirred at reflux (1
h). The mixture was cooled to room temperature, and a black
solid was removed by filtration. The mixture was washed with
ether and acidified with 1 N aqueous HCl, and a solid was
collected by filtration. This was washed with H2O and dried
to yield the desired product. Alternatively, the aqueous layer
resulting after acidification was subjected to an extractive
workup (EtOAc) and concentrated under reduced pressure.
Purification by silica gel flash chromatography provided the
desired product.
P r oced u r e D. To a dry flask at room temperature under
argon were added aryl methyl ketone (1 mmol) in toluene (6

Activity of 3-Aryl-1,3-diketo-Containing Compounds
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15 3191
mL) and NaH (4 mmol) followed by diethyl oxalate (2 mmol),
and the mixture was stirred at 60 °C (3 h). The reaction was
quenched by the addition of H2O, and the mixture was acidified
(to pH 6) with 2 N HCl and subjected to an extractive workup
(EtOAc). Removal of the solvent and purification by silica gel
chromatography provided the desired product.
7.21 (2H, m), 7.11-7.02 (4H, m), 4.40 (2H, q), 1.42 (3H, t);
FABMS (+VE) m/z 313 (M + H)+.
4-(3-Br om op h en yl)-2-h yd r oxy-4-oxob u t -2-en oic Acid
(4a ). Treatment of 3a according to general procedure H
provided 4a as a solid (91% yield): mp 149-150 °C; H-NMR
(DMSO-d6, δ) 8.18 (1H, t, J ) 2 Hz), 8.06 (1H, d, J ) 8 Hz),
7.87 (1H, d, J ) 8 Hz), 7.52 (1H, t, J ) 8 Hz), 7.09 (1H, s);
FABMS (-VE) m/z 269 (M - H)-. Anal. (C10H7BrO4‚0.93H2O)
C, H.
2-H yd r oxy-4-oxo-4-[3-(p h en ylm et h oxy)p h en yl]b u t -2-
en oic Acid (4b). Treatment of 3b according to general
procedure H provided 4b as a solid (30% yield): mp 148-149
°C; H-NMR (DMSO-d6, δ) 7.51-7.65 (1H, d, J ) 8 Hz), 7.62
(1H, d, J ) 2 Hz), 7.51-7.32 (7H, m), 7.06 (1H, s); FABMS
(-VE) m/z 297 (M - H)-. Anal. (C17H14O5) C, H.
2-Hyd r oxy-4-oxo-4-[3-(p h en ylca r bon yla m in o)p h en yl]-
bu t-2-en oic Acid (4c). Treatment of 3c according to general
procedure H provided 4c as a solid (55% yield): mp 184-185
°C; H-NMR (DMSO-d6, δ) 10.48 (1H, s), 8.48 (1H, s), 8.17 (1H,
d, J ) 9 Hz), 7.99 (2H, d, J ) 7 Hz), 7.80 (1H, d, J ) 8 Hz),
7.62-7.53 (4H, m), 7.06 (1H, s); FABMS (-VE) m/z 310 (M -
H)-. Anal. (C17H13NO5‚0.7H2O) C, H, N.
2-Hyd r oxy-4-oxo-(4-p h en oxyp h en yl)bu t-2-en oic Acid
(4d ). Treatment of 3d according to general procedure H
provided 4d as a solid (61% yield): mp 179-180 °C; H-NMR
(DMSO-d6, δ) 8.09 (2H, d, J ) 8.8 Hz), 7.48 (2H, t, J ) 8.8
Hz), 7.27 (1H, t, J ) 7 Hz), 7.16 (2H, d, J ) 8 Hz), 7.08-7.03
(3H, m); FABMS (-VE) m/z 283 (M - H)-. Anal. (C16H12O5‚
0.22H2O) C, H.
2-Hyd r oxy-4-oxo-4-p h en ylbu t-2-en oic Acid (4e). Aceto-
phenone was treated according to general procedure C to
provide 4e which was recrystallized from ether/hexane to
provide 4e as a yellow solid42 (75% yield): mp 155-158 °C;
H-NMR (CDCl3, δ) 7.15 (1H, s), 7.53 (2H, t, J ) 7.3 Hz), 7.65
(1H, t, J ) 7.3 Hz), 8.00 (2H, d, J ) 7.1 Hz); FABMS (+VE)
m/z 191 (M + H)+.
2,4-Dia cetylqu in olin e (5c). Compound 5c was obtained
as a white solid according to literature procedures:43 mp 66-
68 °C (lit.44 69 °C); H-NMR (CDCl3, δ) 2.80 (3H, s), 2.90 (3H,
s), 7.72-7.88 (2H, m), 8.27 (1H, dd, J ) 7.3, 1.2 Hz), 8.39 (1H,
s), 8.57 (1H, dd, J ) 6.8, 0.5 Hz); FABMS (+VE) m/z 214 (M +
H)+.
Eth yl Ester Hyd r olysis. P r oced u r e E. The starting
material was dissolved in dioxane; 1 N aqueous HCl was
added, and the resulting solution was refluxed (4 h). The
solvent was removed under reduced pressure, and H2O was
added to provide a yellow precipitate which was collected,
washed with water, and dried to provide the final product.
P r oced u r e F . The starting material in EtOH was refluxed
(overnight) with KOH (3.8 molar equiv), cooled to room
temperature, diluted with H2O, and adjusted to pH 3 with 3
N aqueous HCl. The resulting precipitate was collected by
filtration and dried to provide the desired product.41
P r oced u r e G. The starting material was dissolved in THF:
EtOH (1:1) and stirred (1 h) with 1 N aqueous NaOH (5 molar
equiv). The reaction mixture was washed twice with ether,
acidified to pH 2 with 2 N aqueous HCl, and subjected to an
exhaustive extractive workup (EtOAc) to provide the desired
product following purification.
P r oced u r e H. A solution of ester (0.3 mmol) in dioxane (3
mL) was stirred with 1 N NaOH (3 mL, 1 h), and then acidified
to pH 4-5 with 2 N aqueous HCl and taken to dryness. Pure
product was produced following preparative HPLC purifica-
tion.
In d ole Acyla tion . P r oced u r e I. To a solution of am-
monium chloride (4 molar equiv) suspended in ethylene
dichloride was added dropwise via syringe acetic anhydride
(2 molar equiv) at room temperature, and the mixture was
stirred at room temperature (15 min). After the mixture had
been stirred for 15 min, indicated indoles (1 molar equiv) in
ethylene dichloride were added dropwise. As the mixture was
stirred at room temperature (2 h), AlCl3 (2 molar equiv) was
added, changing the heterogeneous mixtures to homogeneous
mixtures. Acetic anhydride (1 mol equiv) was added, and the
mixture was stirred (30 min); the mixtures were then poured
into crushed ice and subjected to extractive workups (EtOAc).
The concentration provided residues which were crystallized
from EtOAc.
Rem ova l of P m b F u n ction a lity. P r oced u r e J . A mixture
of Pmb-containing compound, TFA, and ethylenedithiol/H2O
(1:1) was stirred at room temperature (20 h), and then volatiles
were removed under reduced pressure. The residue was
triturated with ether, and the resulting yellow solid was
resuspended in ether, washed with H2O and EtOAc, and dried
to provide the deprotected product.
N-(3-Acetylp h en yl)ben za m id e (2c). To a mixture of 3′-
aminoacetophenone (406 mg, 3 mmol) in pyridine (10 mL) was
added phenylcarbonyl benzoate (814 mg, 3.6 mmol), and the
mixture was stirred at room temperature overnight. After the
reaction was completed, the mixture was poured into ice cold
water (20 mL) and extracted with diethyl ether (20 mL × 3).
The organic layers were combined and dried over Na2SO4.
After removal of the solvent, the residue was chromatographed
(3:1 ratio of hexanes:EtOAc) to provide 2c as a solid (84%
yield): mp 97-99 °C; H-NMR (CDCl3, δ) 8.28 (1H, s), 8.14 (1H,
t, J ) 2 Hz), 8.06 (1H, m), 7.87 (2H, m), 7.67 (1H, dt, J ) 8, 1
Hz), 7.56-7.40 (4H, m), 2.55 (3H, s); FABMS (+VE) m/z 240
(M + H)+.
E t h yl 2-Hyd r oxy-4-oxo-4-[3-(p h en ylca r b on yla m in o)-
p h en yl]bu t-2-en oa te (3c). The compound 2c was treated
according to general procedure D to provide 3c as a solid (99%
yield): mp 100-101 °C; H-NMR (CDCl3, δ) 8.10 (1H, s), 8.07-
8.04 (2H, m), 7.86 (2H, d, J ) 7 Hz), 7.74 (1H, d, J ) 8 Hz),
7.57-7.42 (4H, m), 7.04 (1H, s), 4.37 (2H, q, J ) 8 Hz), 1.39
(3H, t, J ) 8 Hz); FABMS (+VE) m/z 340 (M + H)+. Anal.
(C19H17NO5) C, H, N.
Eth yl 2-Hydr oxy-4-oxo-(4-ph en oxyph en yl)bu t-2-en oate
(3d ). Compound 2d was treated according to general proce-
dure D to provide 3d as an oil (83% yield): H-NMR (CDCl3,
δ) 8.05-7.97 (2H, m), 7.44-7.40 (2H, t, J ) 7.7 Hz), 7.24-
Eth yl 4-[3-(3-Eth oxyca r bon yl-3-h yd r oxyp r op -2-en oyl)-
p h en yl]-2-h yd r oxy-4-oxobu t-2-en oa te (6a ). In a modifica-
tion of general procedure B, diketone 5a (0.50 g, 3.08 mmol)
was coupled with diethyl oxalate (1.80 g, 12.3 mmol) using
NaOEt (0.85 g, 12.5 mmol) in THF (25 mL) to provide a yellow
residue. The residue was suspended in 1 N aqueous HCl,
filtered, washed with H2O, and dried to provide the desired
product 6a as a yellow solid (0.89 g, 80% yield) after crystal-
lization (EtOAc): mp 93-95 °C; H-NMR (DMSO-d6, δ) 1.37
(6H, t, J ) 7.1 Hz), 4.39 (4H, q, J ) 7.1 Hz), 7.21 (2H, s), 7.81
(1H, t, J ) 7.8 Hz), 8.38 (2H, d, J ) 7.6 Hz), 8.59 (1H, s);
FABMS (+VE) m/z 363 (M + H)+. Anal. (C18H18O8‚0.22CH3-
CO2C2H5) C, H.
Eth yl 4-[4-(3-Eth oxyca r bon yl-3-h yd r oxyp r op -2-en oyl)-
p h en yl]-2-h yd r oxy-4-oxobu t-2-en oa te (6b). The treatment
of diketone 5b (1.0 g, 6.17 mmol) as described above for the
preparation of 6a provided the desired product 6b as a shiny
yellow solid (1.83 g, 82% yield) after crystallization (EtOAc):
mp 113-115 °C (lit.45 mp 160 °C); H-NMR (DMSO-d6, δ) 1.37
(6H, t, J ) 7.1 Hz), 4.37 (4H, q, J ) 7.1 Hz), 7.19 (2H, s), 8.25
(4H, s); FABMS (+VE) m/z 363 (M + H)+. Anal. (C18H18O8) C,
H.
Eth yl 4-[4-(3-Eth oxyca r bon yl-3-h yd r oxyp r op -2-en oyl)-
2-qu in olyl]-2-h yd r oxy-4-oxobu t-2-en oa te (6c). Treatment
of diketone 5c as described above for the preparation of 6a
provided the desired product 6c as a solid (77% yield) after
crystallization (EtOAc): mp 148-150 °C; H-NMR (DMSO-d6,
δ) 1.22 (3H, t, J ) 7.3 Hz), 1.33 (3H, t, J ) 7.3 Hz), 4.11 (2H,
q, J ) 6.6 Hz), 4.32 (2H, q, J ) 6.6 Hz), 7.48 (1H, s), 7.73 (1H,
t, J ) 7.3 Hz), 7.88 (1H, t, J ) 7.3 Hz), 8.05 (1H, s), 8.21 (1H,
d, J ) 8.8 Hz), 8.32 (1H, d, J ) 8.1 Hz); FABMS (+VE) m/z
414 (M + H)+.

3192 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15
Pais et al.
4-[3-(3-Ca r boxy-3-h yd r oxyp r op -2-en oyl)p h en yl]-2-h y-
d r oxy-4-oxobu t-2-en oic Acid (7a ). Treatment of diester 6a
according to general procedure F gave a yellow solid. Recrys-
tallization (AcOH) provided diacid 7a as beige solid (75%
yield): mp >250 °C; H-NMR (DMSO-d6, δ) 7.01 (2H, s), 7.76
(1H, t, J ) 8.1 Hz), 8.31 (2H, d, J ) 7.6 Hz), 8.56 (1H, s);
FABMS (-VE) m/z 305 (M - H)-. HRMS calcd for C14H9O8:
305.0297. Found: 305.0303.
4-[4-(3-Ca r boxy-3-h yd r oxyp r op -2-en oyl)p h en yl]-2-h y-
d r oxy-4-oxobu t-2-en oic Acid (7b). Treatment of diester 6b
as described in general procedure F and crystallization of the
product from EtOAc provided 7b as a yellow solid (78%
yield): mp 249-250 °C; H-NMR (DMSO-d6, δ) 7.17 (2H, s),
8.23 (4H, s); FABMS (-VE) m/z 306 (M•)-.
4-[4-(3-Ca r boxy-3-h yd r oxyp r op -2-en oyl)-2-qu in olyl]-2-
h yd r oxy-4-oxobu t-2-en oic Acid (7c). Treatment of diester
6c as described in general procedure G provided a brown solid
in 65% yield by HPLC: H-NMR (DMSO-d6, δ) 6.69 (2H, s),
7.85 (1H, t, J ) 8.1 Hz), 7.97 (1H, t, J ) 6.6 Hz), 8.13 (1H, s),
8.29 (2H, t, J ) 8.1 Hz); FABMS (-VE) m/z 356 (M - H)-.
Anal. (C17H11NO8‚0.84H2O) C, H, N.
3-Acetyl-5-ch lor oin d ole (8b). Compound 8b was prepared
according to general procedure I and was obtained as a white
solid in 75% yield: H-NMR (DMSO-d6, δ) 2.45 (3H, s), 7.22
(1H, dd, J ) 8.1, 2.2 Hz), 7.49 (1H, d, J ) 8.8 Hz), 8.14 (1H,
s), 8.38 (1H, d, J ) 2.9 Hz), 12.10 (1H, brs); FABMS (+VE)
m/z 194 (M + H)+.
4-Acetylqu in olin e (8c). Compound 8c was obtained as a
white solid according to literature procedures:43 mp 50-51 °C;
H-NMR (CDCl3, δ) 2.83 (3H, s), 7.61-7.84 (2H, m), 7.87 (1H,
d, J ) 8.1 Hz), 8.10-8.31 (3H, m); FABMS (+VE) m/z 172 (M
+ H)+.
3-Acetyl-5-flu or oin d ole (8f). Compound 8f was prepared
according to general procedure I and was obtained as a white
solid in 73% yield: mp 200-201 °C (lit.46 200-201.5 °C);
H-NMR (DMSO-d6, δ) 2.48 (3H, s), 7.11 (1H, td, J ) 6.1, 2.4
Hz), 7.52 (1H, dd, J ) 4.6, 4.2 Hz), 7.86 (1H, dd, J ) 10, 2.2
Hz), 8.42 (1H, d, J ) 2.9 Hz), 12.06 (1H, brs); FABMS (+VE)
m/z 178 (M + H)+.
H-NMR (DMSO-d6, δ) 1.33 (3H, t, J ) 6.8 Hz), 4.27 (2H, q, J
) 7.1 Hz), 7.22 (1H, s), 7.73 (1H, t, J ) 8.5 Hz), 7.88 (1H, t, J
) 8.5 Hz), 8.08 (1H, d, J ) 7.6 Hz), 8.18 (2H, d, J ) 8.1 Hz),
8.53 (1H, d, J ) 7.6 Hz); FABMS (+VE) m/z 272 (M + H)+.
E t h yl 4-(2-F er r ocen yl)-2-h yd r oxy-4-oxob u t -2-en oa t e
(9d ). Treatment of acetylferrocene as described in general
procedure C and purification of the product by silica gel flash
chromatography (20:80 ratio of EtOAc:hexane) gave pure 9d
as a violet solid (60% yield): mp 66-69 °C; H-NMR (CDCl3,
δ) 1.42 (3H, t, J ) 7.1 Hz), 4.26 (5H, s), 4.39 (2H, q, J ) 7.1
Hz), 4.68 (2H, s), 4.92 (2H, s), 6.56 (1H, s); FABMS (+VE) m/z
328 (M + H)+.
Eth yl 4-[(3,5-Diben zyloxy)p h en -1-yl]-2-h yd r oxy-4-oxo-
bu t-2-en oa te (9e). Treatment of 3,5-dibenzyloxyacetophenone
(8e) as described in general procedure B and crystallization
of the product from EtOAc/hexane gave 9e as a bright yellow
solid: mp 77-78 °C; H-NMR (CDCl3, δ) 1.51 (3H, t, J ) 7.1
Hz), 4.49 (2H, q, J ) 7.1 Hz), 5.18 (4H, s), 6.95 (1H, s), 7.11
(1H, s), 7.33 (2H, s), 7.35-7.59 (10H, m); FABMS (+VE) m/z
433 (M + H)+. Anal. (C26H24O6‚0.026CH3CO2C2H5) C, H.
4-(In d ol-3-yl)-2-h yd r oxy-4-oxobu t-2-en oic Acid (10a ).
Treatment of ester 9a as described in general procedure E and
crystallization from EtOAc provided 9a as a yellow solid (56%
yield):25 mp 202-210 °C; H-NMR (DMSO-d6, δ) 7.06 (1H, s),
7.30 (2H, t, J ) 3.6 Hz), 7.53 (1H, s), 8.27 (1H, s), 8.75 (1H, d,
J ) 2.7 Hz), 12.44 (1H, s); FABMS (-VE) m/z 230 (M - H)-.
4-(5-Ch lor oin dol-3-yl)-2-h ydr oxy-4-oxobu t-2-en oic Acid
(10b). Treatment of ester 9b as described in general procedure
E provided 10b as a brown solid (65% yield): mp 220-225 °C
(lit.24 220-225 °C); H-NMR (DMSO-d6, δ) 7.06 (1H, s), 7.29
(1H, dd, J ) 8.7, 2.4 Hz), 7.53 (1H, d, J ) 8.7 Hz), 8.21 (1H,
d, J ) 2.4 Hz), 8.77 (1H, d, J ) 3.6 Hz), 12.5 (1H, brm).
2-Hydr oxy-4-oxo-4-(qu in olin -4-yl)bu t-2-en oic Acid (10c).
Treatment of ester 9c as described in general procedure G and
purification by HPLC provided 10c as a yellow solid (75%
yield): H-NMR (DMSO-d6, δ) 7.74 (1H, t, J ) 8.1 Hz), 7.86
(1H, t, J ) 8.8 Hz), 8.08 (1H, d, J ) 9.5 Hz), 8.14 (2H, t, J )
8.8 Hz), 8.55 (1H, d, J ) 8.8 Hz); FABMS (-VE) m/z 242 (M -
H)-.
4-(2-Fer r ocen yl)-2-h ydr oxy-4-oxobu t-2-en oic Acid (10d).
Treatment of ester 9d as described in general procedure G and
purification by silica gel flash chromatography (90:10 ratio of
CHCl3:MeOH) provided 10b as a violet solid (60% yield): mp
160-163 °C; H-NMR (CDCl3, δ) 4.23 (5H, s), 4.69 (2H, s), 4.92
(2H, s), 7.27 (1H, s); FABMS (-VE) m/z 299 (M - H)-. HR-
FABMS calcd for C14H11FeO4 (M - H): 299.00067. Found:
299.00158.
4-[(3,5-Diben zyloxy)p h en -1-yl]-2-h yd r oxy-4-oxobu t-2-
en oic Acid (10e). Treatment of ester 9e as described in
general procedure E and crystallization from EtOAc provided
10e as a yellow solid (75% yield): mp 170-172 °C; H-NMR
(DMSO-d6, δ) 5.15 (4H, s), 6.95 (1H, t, J ) 2.3 Hz), 7.03 (1H,
s), 7.21 (2H, d, J ) 2.3 Hz), 7.35-7.61 (10H, m); FABMS (-VE)
m/z 403 (M - H)-. Anal. (C24H20O6‚0.15CH3CO2C2H5) C, H.
Eth yl 1-(4-Meth oxyben zyl)tetr azole-5-car boxylate (11).
By using literature procedures,26 compound 11 was obtained
as a white solid in 58% yield: mp 51-53 °C (lit.26 50-52 °C);
H-NMR (CDCl3, δ) 1.39 (3H, t, J ) 7.1 Hz), 3.85 (3H, s), 4.47
(2H, q, J ) 7.1 Hz), 5.80 (2H, s), 6.81 (2H, d, J ) 8.8 Hz), 7.42
(2H, d, J ) 8.8 Hz).
1-(In d ol-3-yl)-3-h yd r oxy-3-[1-(4-m eth oxyben zyl)tetr a -
zol-5-yl]p r op en on e (12a ). Coupling of 8a with 11 as indi-
cated in general procedure A and crystallization from EtOAc
provided 12a as a yellow solid (88% yield): mp 210-215 °C;
H-NMR (DMSO-d6, δ) 3.69 (3H, s), 6.88 (2H, d, J ) 10.3 Hz),
7.20 (1H, s), 7.20-7.27 (2H, m), 7.28 (2H, d, J ) 10.5 Hz),
7.45-7.51 (1H, m), 8.09-8.15 (1H, m), 8.64 (1H, d, J ) 5.2
Hz); FABMS (+VE) m/z 376 (M + H)+.
1-(5-Ch lor oin d ol-3-yl)-3-h yd r oxy-3-[1-(4-m eth oxyben -
zyl)tetr a zol-5-yl]p r op en on e (12b). Coupling of 8b with 11
as indicated in general procedure A and crystallization from
EtOAc to provide 12b as a yellow solid (84% yield): mp 200-
205 °C; H-NMR (DMSO-d6, δ) 3.72 (3H, s), 5.94 (2H, s), 6.96
(2H, d, J ) 7.3 Hz), 7.28 (1H, s), 7.36 (3H, d, J ) 8.8 Hz), 7.53
Ben zyl 3-Acetylin d ole-5-ca r boxyla te (8g). A mixture of
indole-5-carboxylic acid (0.20 g, 1.24 mmol), benzyl bromide
(0.89 mL, 7.45 mmol), and NaHCO3 (0.63 g, 7.45 mmol) in
DMF (5 mL) was stirred at room temperature (3 days). The
mixture was poured into H2O and subjected to an extractive
workup (EtOAc), and the residue crystallized in EtOAc to
provide benzyl indole-5-carboxylate as a colorless solid (0.28
g, 90% yield):47 mp 128-130 °C (lit.48 127-129 °C); H-NMR
(CDCl3, δ) 5.49 (2H, s), 6.73 (1H, d, J ) 2.2 Hz), 7.33-7.60
(6H, m), 8.05 (1H, dd, J ) 7.1, 1.5 Hz), 8.57 (2H, s). Acylation
as indicated in general procedure I provided 8g as a colorless
solid in 65% yield: mp 217-219 °C; H-NMR (DMSO-d6, δ) 2.55
(3H, s), 5.36 (2H, s), 7.30-7.60 (6H, m), 7.86 (1H, dd, J ) 6.6,
2.2 Hz), 8.45 (1H, s), 8.89 (1H, d, J ) 1.5 Hz), 12.31 (1H, brs);
FABMS (+VE, NBA) m/z 294 (M + H)+. Anal. (C18H15NO3‚
0.002H2O) C, H, N.
Eth yl 4-(In d ol-3-yl)-2-h yd r oxy-4-oxobu t-2-en oa te (9a ).
Treatment of 3-acetyl indole (8a ) as indicated in general
procedure B with recrystallization from MeOH/hexane gave
9a as a yellow solid (80% yield): mp >300 °C; H-NMR (DMSO-
d6, δ) 1.35 (3H, t, J ) 7.1 Hz), 4.31 (2H, q, J ) 7.1 Hz), 6.97
(1H, s), 7.28 (2H, t, J ) 3.6 Hz), 7.53 (1H, d, J ) 6.4 Hz), 8.28
(1H, d, J ) 7.1 Hz), 8.67 (1H, s); FABMS (+VE) m/z 260 (M +
H)+. Anal. (C14H13NO4) C, H, N.
Eth yl 4-(5-Ch lor oin d ol-3-yl)-2-h yd r oxy-4-oxobu t-2-en o-
a te (9b). Treatment of 3-acetyl-5-chloroindole as indicated in
general procedure B with recrystallization of the product from
dioxane gave 9b as a yellow solid (82% yield): mp 219-225
°C; H-NMR (DMSO-d6, δ) 1.36 (3H, t, J ) 7.1 Hz), 4.35 (2H,
q, J ) 7.1 Hz), 7.08 (1H, s), 7.35 (1H, d, J ) 8.5 Hz), 7.75 (1H,
d, J ) 8.6 Hz), 8.25 (1H, s), 8.87 (1H, d, J ) 1.2 Hz), 12.67
(1H, brs); FABMS (+VE) m/z 294 (M + H)+.
Eth yl 4-(Qu in ol-4-yl)-2-h ydr oxy-4-oxobu t-2-en oate (9c).
Treatment of 4-acetylquinoline according to general procedure
B gave 9c as a brown solid (86% yield): mp 145-150 °C;

Activity of 3-Aryl-1,3-diketo-Containing Compounds
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15 3193
(1H, d, J ) 8.1 Hz), 8.25 (1H, s), 8.58 (1H, s), 12.38 (1H, brs);
indicated in general procedure A and crystallization from
FABMS (+VE) m/z 410 (M + H)+. HR-FABMS calcd for C20H15
ClN5O3 (M - H): 408.0863. Found: 408.0878.
-
EtOAc provided 15a as a yellow solid (85%, yield): mp 217-
218 °C; H-NMR (DMSO-d6
, δ) 3.75 (6H, s), 5.98 (2H, s), 6.48-
6.55 (3H, m), 7.27-7.38 (3H, m), 7.53-7.61 (1H, m), 8.18-
8.25 (1H, m), 8.82 (1H, s), 12.6 (1H, s); FAB-MS (+VE) m/z
406 (M + H)+. HR-FABMS calcd for C21H18N5O4 (M - H):
404.1359. Found: 404.1367.
1-(5-F lu or oin d ol-3-yl)-3-h yd r oxy-3-[1-(4-m eth oxylben -
zyl)tetr a zol-5-yl]p r op en on e (12f). Coupling of 8f with 11
as indicated in general procedure A provided 12f as a yellow
solid (60% yield): mp 212-215 °C; H-NMR (DMSO-d6, δ) 3.76
(3H, s), 5.97 (2H, s), 6.97 (2H, d, J ) 8.3 Hz), 7.19 (1H, t, J )
9.5 Hz), 7.28 (1H, s), 7.38 (2H, d, J ) 8.5 Hz), 7.58 (1H, dd, J
) 3.4, 4.4 Hz), 7.90 (1H, d, J ) 8.8 Hz), 8.89 (1H, s), 12.71
(1H, brs); FABMS (+VE) m/z 394 (M + H)+. HR-FABMS calcd
for C20H15FN5O3 (M - H): 392.1159. Found: 392.1143.
1-[(5-Ben zyloxyca r bon yl)in d ol-3-yl]-3-h yd r oxy-3-[1-(4-
m eth oxyben zyl)tetr a zol-5-yl]p r op en on e (12g). Coupling
of 8g with 11 as indicated in general procedure A and
crystallization from EtOAc provided 12g as a yellow solid (75%
yield): mp 195-198 °C; H-NMR (DMSO-d6, δ) 3.47 (3H, s, J
) 3 Hz), 5.32 (2H, s), 5.82 (2H, s), 6.40 (2H, d, J ) 8.5 Hz),
6.91 (1H, s), 7.28-7.56 (8H, m), 7.87 (1H, dd, J ) 6.8, 1.7 Hz),
8.39 (1H, s), 9.38 (1H, s), 12.16 (1H, brs); FABMS (+VE, NBA)
m/z 510 (M + H)+. HR-FABMS calcd for C28H22N5O5 (M - H):
508.1621. Found: 508.1627.
1-(In d ol-3-yl)-3-h yd r oxy-3-(1H -t e t r a zol-5-yl)p r op e -
n on e (13a ). Treatment of 12a as described in general proce-
dure J provided a yellow solid which was recrystallized from
DMF to provide 13a as a yellow solid (75% yield): mp 235-
240 °C; H-NMR (DMSO-d6, δ) 7.25 (1H, s), 7.26-7.31 (2H, m),
7.51-7.55 (1H, m), 8.22 (1H, dd, J ) 5.9, 2.9 Hz), 8.76 (1H, d,
J ) 3.7 Hz), 12.45 (1H, s); CIMS (NH3) m/z 273 (M + NH4)+.
Anal. (C12H9N5O2‚0.86C3H7NO) C, H, N.
1-(5-Ch lor oin d ol-3-yl)-3-h yd r oxy-3-(1H -t et r a zol-5-yl)-
p r op en on e (13b). Treatment of 12b as described in general
procedure J provided 13b as a yellow solid (70% yield): mp
250 °C; H-NMR (DMSO-d6, δ) 7.26 (1H, s), 7.32 (1H, dd, J )
8.7, 2.1 Hz), 7.56 (1H, d, J ) 8.7 Hz), 8.21 (1H, d, J ) 2.1 Hz),
8.84 (1H, d, J ) 3.3 Hz), 12.6 (1H, brs); FABMS (-VE) m/z
288 (M - H)+.
1-(5-F lu or oin d ol-3-yl)-3-h yd r oxy-3-(1H -t et r a zol-5-yl)-
p r op en on e (13f). Treatment of 12f as described in general
procedure J provided 13f as a yellow solid (86% yield): mp
260 °C dec; H-NMR (DMSO-d6, δ) 7.15-7.31 (2H, m), 7.55-
7.58 (1H, m), 7.89-7.99 (1H, m), 8.89 (1H, t, J ) 3.2 Hz), 12.63
(1H, brs); FABMS (+VE) m/z 274 (M + H)+.
1-[(3,5-Diben zyloxy)p h en -1-yl]-3-h yd r oxy-3-[1-(4-m eth -
oxyben zyl)tetr a zol-5-yl]p r op en on e (16b). Coupling of 8e
with 11 as indicated in general procedure A provided 16b as
a yellow solid (60% yield): mp 118-120 °C; H-NMR (CDCl3,
δ) 3.78 (3H, s), 5.09 (4H, s), 5.94 (2H, s), 6.85 (1H, s), 6.89
(1H, s), 7.23 (2H, s), 7.31-7.49 (10H, m); FABMS (+VE) m/z
549 (M + H)+.
Refer en ces
(1) A preliminary account of this information has been presented
in: Burke, T. R., J r.; Pais, C. G.; Zhang, X.; Marchand, C.;
Neamati, N.; Pommier, Y.; Svarovskaia, E. S.; Pathak, V. K.
Structure-activity investigation of 3-aryl-1,3-diketo containing
compounds as HIV-1 integrase inhibitors. Second HIV DRP
Symposium: Antiviral Drug Resistance, Chantilly, VA, 2001.
(2) Palella, F. J ., J r.; Delaney, K. M.; Moorman, A. C.; Loveless, M.
O.; Fuhrer, J .; Satten, G. A.; Aschman, D. J .; Holmberg, S. D.
Declining morbidity and mortality among patients with ad-
vanced human immunodeficiency virus infection. HIV Outpa-
tient Study Investigators. N. Engl. J . Med. 1998, 338, 853-860.
(3) Richman, D. D. HIV chemotherapy. Nature 2001, 410, 995-
1001.
(4) LaFemina, R. L.; Schneider, C. L.; Robbins, H. L.; Callahan, P.
L.; LeGrow, K.; Roth, E.; Schleif, W. A.; Emini, E. A. Require-
ment of active human immunodeficiency virus type 1 integrase
for productive infection of human T-lymphoid cells. J . Virol.
1992, 66, 7414-7419.
(5) Pommier, Y.; Neamati, N. Inhibitors of human immunodeficiency
virus integrase. Adv. Virus Res. 1999, 52, 427-458.
(6) Farnet, C. M.; Wang, B. B.; Lipford, J . R.; Bushman, F. D.
Differential inhibition of HIV-1 preintegration complexes and
purified integrase protein by small molecules. Proc. Natl. Acad.
Sci. U.S.A. 1996, 93, 9742-9747.
(7) Pommier, Y.; Marchand, C.; Neamati, N. Retroviral integrase
inhibitors year 2000: update and perspectives. Antiviral Res.
2000, 47, 139-148.
(8) Fujishita, T.; Yoshinaga, T.; Sato, A. (Shionogi & Co., Ltd.).
Preparation of aromatic heterocycle compounds having HIV
integrase inhibiting activities. PCT Int. Appl. WO-00039086,
2000; pp 554.
(9) Uenaka, M.; Kawata, K.; Nagai, M.; Endoh, T. (Shionogi & Co.,
Ltd.). Novel processes for the preparation of substituted prope-
none derivatives. PCT Int. Appl. WO-00075122, 2000; pp 97.
(10) Selnick, H. G.; Hazuda, D. J .; Egbertson, M.; Guare, J . P., J r.;
Wai, J . S.; Young, S. D.; Clark, D. L.; Medina, J . C. (Merck and
Co., Inc.). Preparation of nitrogen-containing 4-heteroaryl-2,4-
dioxobutyric acids useful as HIV integrase inhibitors. PCT Int.
Appl. WO-09962513, 1999; pp 287.
(11) Young, S. D.; Egbertson, M.; Payne, L. S.; Wai, J . S.; Fisher, T.
E.; Guare, J . P., J r.; Embrey, M. W.; Tran, L.; Zhuang, L.; Vacca,
J . P.; Langford, M.; Melamed, J .; Clark, D. L.; Medina, J . C.;
J aen, J . (Merck and Co., Inc.). Preparation of aromatic and
heteroaromatic 4-aryl-2,4-dioxobutyric acid derivatives useful as
HIV integrase inhibitors. PCT Int. Appl. WO-9962520, 1999; pp
319.
(12) Payne, L. S.; Tran, L. O.; Zhuang, L. H.; Young, S. D.; Egbertson,
M. S.; Wai, J . S.; Embrey, M. W.; Fisher, T. E.; Guare, J . P.;
Langford, H. M.; Melamed, J . Y.; Clark, D. L. Preparation of
1,3-diaryl-1,3-propanediones as HIV integrase inhibitors. PCT
Int. Appl. WO-0100578 (Merck & Co., Inc.; Tularik, Inc.). USA,
2001; pp 236.
(13) Goldgur, Y.; Craigie, R.; Cohen, G. H.; Fujiwara, T.; Yoshinaga,
T.; Fujishita, T.; Sugimoto, H.; Endo, T.; Murai, H.; Davies, D.
R. Structure of the HIV-1 integrase catalytic domain complexed
with an inhibitor: A platform for antiviral drug design. Proc.
Natl. Acad. Sci. U.S.A. 1999, 96, 13040-13043.
(14) Espeseth, A. S.; Felock, P.; Wolfe, A.; Witmer, M.; Grobler, J .;
Anthony, N.; Egbertson, M.; Melamed, J . Y.; Young, S.; Hamill,
T.; Cole, J . L.; Hazuda, D. J . HIV-1 integrase inhibitors that
compete with the target DNA substrate define a unique strand
transfer conformation for integrase. Proc. Natl. Acad. Sci. U.S.A.
2000, 97, 11244-11249.
(15) Pluymers, W.; Neamati, N.; Pannecouque, C.; Fikkert, V.;
Marchand, C.; Burke, T. R.; Pommier, Y.; Schols, D.; DeClercq,
E.; Debyser, Z.; Witvrouw, M. Viral entry as the primary target
for the anti-HIV activity of chicoric acid and its tetra-acetyl
esters. Mol. Pharmacol. 2000, 58, 641-648.
1-(5-Ca r boxyin d ol-3-yl)-3-h yd r oxy-3-(1H-tetr a zol-5-yl)-
p r op en on e (13g). A solution of 12g (0.05 g, 0.098 mmol) in
THF (2 mL) with Pd‚C (100 mg) was stirred at room temper-
ature under H2 supplied by a balloon (4 h). The mixture was
filtered through Celite, and the filter pad was washed thor-
oughly with THF. The combined organic layer was dried,
concentrated under reduced pressure, and then treated as
indicated in general procedure J to provide the desired product
13g as a yellow solid (10 mg, 34% yield): H-NMR (DMSO-d6,
δ) 7.28 (1H, s), 7.62 (1H, dd, J ) 8.3, 0.98 Hz), 7.92 (1H, d, J
) 12.5 Hz), 8.85 (1H, s), 8.95 (1H, s), 12.68 (1H, brs); FABMS
(-VE, NBA) m/z 298 (M - H)-. Anal. (C13H9N5O4) C, H, N.
Eth yl 1-(3,5-Dim eth oxyben zyl)tetr a zole-5-ca r boxyla te
(14). A mixture of 3,5-dimethoxybenzyl bromide (1.0 g, 4.33
mmol) and sodium azide (0.338 g, 5.19 mmol) in 70% aqueous
EtOH (5 mL) was stirred at 58 °C (overnight). The mixture
was cooled to room temperature, diluted with H2O, and
subjected to an extractive workup (CHCl3). Concentration and
purification by silica gel flash chromatography (95:5 ratio of
hexane:EtOAc) provided the intermediate 3,5-dimethoxybenzyl
azide as a colorless oil (0.68 g, 81% yield): H-NMR (CDCl3, δ)
3.80 (6H, s), 4.27 (2H, s), 6.43 (1H, t, J ) 2.2 Hz), 6.46 (2H, d,
J ) 2.2 Hz). Treatment of 3,5-dimethoxybenzyl azide in a
manner similar to that used to prepare compound 11 provided
14 as a white solid in 70% yield: mp 95-97 °C; H-NMR
(CDCl3, δ) 1.41 (3H, t, J ) 7.1 Hz), 3.72 (6H, s), 4.47 (2H, q, J
) 7.1 Hz), 5.8 (2H, s), 6.37 (1H, t, J ) 2.2 Hz), 6.44 (2H, d, J
) 2.2 Hz); FABMS (+VE) m/z 293 (M + H)+. Anal. (C13H16N4O4)
C, H, N.
1-(In d ol-3-yl)-3-h yd r oxy-3-[1-(3,5-d im eth oxyben zyl)tet-
r a zol-5-yl]p r op en on e (15a ). Coupling of 8a with 14 as

3194 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 15
Pais et al.
(34) Gaussian 98; G. Gaussian Inc.: Pittsburgh, PA, 1998.
(35) Rarey, M.; Kramer, B.; Lengauer, T.; Klebe, G. A fast flexible
docking method using an incremental construction algorithm.
J . Mol. Biol. 1996, 261, 470-489.
(36) Brooks, B. R.; Bruccoleri, R. E.; Olafson, B. D.; States, D. J .;
Swaminathan, S.; Karplus, M. Charmm - a Program for
Macromolecular Energy, Minimization, and Dynamics Calcula-
tions. J . Comput. Chem. 1983, 4, 187-217.
(37) MacKerell, A. D.; Bashford, D.; Bellott, M.; Dunbrack, R. L.;
Evanseck, J . D.; Field, M. J .; Fischer, S.; Gao, J .; Guo, H.; Ha,
S.; J oseph-McCarthy, D.; Kuchnir, L.; Kuczera, K.; Lau, F. T.
K.; Mattos, C.; Michnick, S.; Ngo, T.; Nguyen, D. T.; Prodhom,
B.; Reiher, W. E.; Roux, B.; Schlenkrich, M.; Smith, J . C.; Stote,
R.; Straub, J .; Watanabe, M.; Wiorkiewicz-Kuczera, J .; Yin, D.;
Karplus, M. All-atom empirical potential for molecular modeling
and dynamics studies of proteins. J . Phys. Chem. B 1998, 102,
3586-3616.
(38) Foloppe, N.; MacKerell, A. D. All-atom empirical force field for
nucleic acids: I. Parameter optimization based on small molecule
and condensed phase macromolecular target data. J . Comput.
Chem. 2000, 21, 86-104.
(39) This study utilized the high-performance computational capa-
bilities of the SGI Origin 2000 system and the Biowulf/LoBoS3
cluster at the Center for Information Technology, National
Institutes of Health, Bethesda, MD.
(16) Vandegraaff, N.; Kumar, R.; Hocking, H.; Burke, T. R., J r.; Mills,
J .; Rhodes, D.; Burrell, C. J .; Li, P. Specific inhibition of human
immunodeficiency virus type
1 (HIV-1) integration in cell
culture: putative inhibitors of HIV-1 integrase. Antimicrob.
Agents Chemother. 2001, 45, 2510-2516.
(17) Pluymers, W.; Pais, G.; Neamati, N.; Pannecouque, C.; Fikkert,
V.; Pommier, Y.; Clercq, E. D.; Witvrouw, M.; Burke, T. R., J r.;
Debyser, Z. Inhibition of HIV-1 integrase activity and HIV-1
replication by a series of diketo derivatives. Mol. Pharmacol.,
in review.
(18) Hazuda, D. J .; Felock, P.; Witmer, M.; Wolfe, A.; Stillmock, K.;
Grobler, J . A.; Espeseth, A.; Gabryelski, L.; Schleif, W.; Blau,
C.; Miller, M. D. Inhibitors of strand transfer that prevent
integration and inhibit HIV-1 replication in cells. Science 2000,
287, 646-650.
(19) Marchand, C.; Zhang, X.; Pais, G. C. G.; Cowansage, K.; Neamati,
N.; Burke, T. R., J r.; Pommier, Y. Structural determinants for
HIV-1 integrase inhibition by â-diketo acids. J . Biol. Chem. 2002,
277, 12596-12603.
(20) Neamati, N. Structure-based HIV-1 integrase inhibitor design:
a future perspective. Expert Opin. Invest. Drugs 2001, 10, 281-
296.
(21) Neamati, N.; Marchand, C.; Winslow, H. E.; Pommier, Y. Human
immunodeficiency virus type
1 integrase-targeted inhibitor
design. Antiretroviral Ther. 2001, 87-104.
(22) Young, S. D. Inhibition of HIV-1 integrase by small molecules:
the potential for a new class of AIDS chemotherapeutics. Curr.
Opin. Drug Discovery Dev. 2001, 4, 402-410.
(40) Quanta/ CHARMM 2000 and InsightII 2000; Accelrys Inc.: San
Diego, CA, 2000.
(41) Flynn, D. L.; Belliotti, T. R.; Boctor, A. M.; Connor, D. T.;
Kostlan, C. R.; Nies, D. E.; Ortwine, D. F.; Schrier, D. J .; Sircar,
J . J . Styrylpyrazoles, styrylisoxazoles, and styrylisothiazoles -
novel 5 lipoxygenase and cyclooxygenase inhibitors. J . Med.
Chem. 1991, 34, 518-525.
(42) Andreichikov, Y. S.; Maslivets, A. N.; Smirnova, L. I.; Krasnykh,
O. P.; Kozlov, A. P.; Perevozchikov, L. A. Five-member 2,3-
dioxoheterocycles. 5. Synthesis of 1-aryl-4-aroyl-5-methoxycar-
bonyl-2,3-dihydro-2,3-pyrrolediones - Their interaction with
water and alcohols. Zh. Org. Khim. 1987, 23, 1534-1543.
(43) Fontana, F.; Minisci, F.; Barbosa, M. C. N.; Vismara, E.
Homolytic acylation of protonated pyridines and pyrazines with
alpha-keto acids - the problem of monoacylation. J . Org. Chem.
1991, 56, 2866-2869.
(44) Caronna, T.; Fronza, G.; Minisci, F.; Porta, O. Homolytic
acylation of protonated pyridine and pyrazine derivatives. J .
Chem. Soc., Perkin Trans. 2 1972, 2035-2038.
(45) El-Bahaie, S.; Assy, M. G.; Hassanien, M. M. Synthesis and
biological activity of some azoles and azines. J . Indian Chem.
Soc. 1990, 67, 757-758.
(46) Ketcha, D. M.; Gribble, G. W. A convenient synthesis of 3-acylin-
doles via friedel-crafts acylation of 1-(phenylsulfonyl)indole -
A new route to pyridocarbazole-5,11-quinones and ellipticine.
J . Org. Chem. 1985, 50, 5451-5457.
(47) Boger, D. L.; Ledeboer, M. W.; Kume, M.; Searcey, M.; J in, Q.
Total synthesis and comparative evaluation of luzopeptin A-C
and quinoxapeptin A-C. J . Am. Chem. Soc. 1999, 121, 11375-
11383.
(48) J acobs, R. T.; Brown, F. J .; Cronk, L. A.; Aharony, D.; Buckner,
C. K.; Kusner, E. J .; Kirkland, K. M.; Neilson, K. L. Substituted
3-(phenylmethyl)-1H-indole-5-carboxamides and 1-(phenyl-
methyl)indole-6-carboxamides as potent, selective, orally active
antagonists of the peptidoleukotrienes. J . Med. Chem. 1993, 36,
394-409.
(23) Wai, J . S.; Egbertson, M. S.; Payne, L. S.; Fisher, T. E.; Embrey,
M. W.; Tran, L. O.; Melamed, J . Y.; Langford, H. M.; Guare, J .
P., J r.; Zhuang, L.; Grey, V. E.; Vacc, J . P.; Holloway, M. K.;
Naylor-Olsen, A. M.; Hazuda, D. J .; Felock, P. J .; Wolfe, A. L.;
Stillmock, K. A.; Schleif, W. A.; Gabryelski, L. J .; Young, S. D.
4-Aryl-2,4-dioxobutanoic acid inhibitors of HIV-1 integrase and
viral replication in cells. J . Med. Chem. 2000, 43, 4923-4926.
(24) Fujishita, T.; Yoshinaga, T.; Yamauchi, H. (Shionogi & Co., Ltd.).
Indole derivatives with antiviral activity. PCT Int. Appl. WO-
9950245, 1999.
(25) Barrett, C. B.; Beer, R. J . S.; Dodd, G. M.; Robertson, A. The
chemistry of Bacteria. Part VI. The synthesis of a trimethyl
derivative of the C20 acid from Violacein. J . Chem. Soc. 1957,
4810-4813.
(26) Klaubert, D. H.; Sellstedt, J . H.; Guinosso, C. J .; Bell, S. C.;
Capetola, R. J . 5-Tetrazolecarboxamides and their salts: new
orally active antiallergy agents. J . Med. Chem. 1981, 24, 748-
752.
(27) Tang, Y.; Nicklaus, M. C. Refined model of the HIV-1 integrase-
viral DNA complex used for inhibitor docking studies. Presented
at the 222nd ACS National Meeting of the American Chemical
Society, Chicago, IL, Aug 26-30, 2001.
(28) Brautigam, C. A.; Steitz, T. A. Structural and functional insights
provided by crystal structures of DNA polymerases and their
substrate complexes. Curr. Opin. Struct. Biol. 1998, 8, 54-63.
(29) Thornber, C. W. Isosterism and molecular modification in drug
design. Chem. Soc. Rev. 1979, 8, 563-580.
(30) Patani, G. A.; LaVoie, E. J . Bioisosterism: A rational approach
in drug design. Chem. Rev. 1996, 96, 3147-3176.
(31) Marchand, C.; Neamati, N.; Pommier, Y. In vitro human
immunodeficiency virus type 1 integrase assays. Methods En-
zymol. 2001, 340, 624-633.
(32) SYBYL 6.7.1; S. Tripos Inc.: St. Louis, MO, 2001.
(33) Tang, Y.; Nicklaus, M. C. Molecular modeling of full-length
HIV-1 integrase and its complexes with viral and human DNA.
Presented at the 221st ACS National Meeting of the American
Chemical Society, San Diego, CA, Apr 1-5, 2001.
J M020037P