Synthesis and biological evaluation of certain alkenyldiarylmethanes as anti-HIV-1 agents which act as non-nucleoside reverse transcriptase inhibitors

Article abstract of DOI:10.1021/jm960082v

Several novel alkenyldiarylmethane (ADAM) non-nucleoside HIV-1 reverse transcriptase inhibitors were synthesized. The most potent of these proved to be 3',3'-dibromo-4',4'-dimethoxy-5',5'-bis(methoxycarbonyl)-1,1-diphenyl-1- heptene (8). ADAM 8 inhibited

Full text of DOI:10.1021/jm960082v

J . Med. Chem. 1996, 39, 3217-3227
3217
Syn th esis a n d Biologica l Eva lu a tion of Cer ta in Alk en yld ia r ylm eth a n es a s
An ti-HIV-1 Agen ts Wh ich Act a s Non -Nu cleosid e Rever se Tr a n scr ip ta se
In h ibitor s
Mark Cushman,*,† W. Marek Golebiewski,† Lisa Graham,‡ J im A. Turpin,‡ William G. Rice,‡
Valerie Fliakas-Boltz,§ and Robert W. Buckheit, J r.§
Department of Medicinal Chemistry and Molecular Pharmacology, School of Pharmacy and Pharmacal Sciences,
Purdue University, West Lafayette, Indiana 47907, Laboratory of Antiviral Drug Mechanisms, SAIC Frederick,
National Cancer Institute-Frederick Cancer Research and Development Center, Frederick, Maryland 21702-1201, and
Virology Research Group, Southern Research Institute-Frederick Research Center, Frederick, Maryland 21701
Received J anuary 26, 1996X
Several novel alkenyldiarylmethane (ADAM) non-nucleoside HIV-1 reverse transcriptase
inhibitors were synthesized. The most potent of these proved to be 3′,3′′-dibromo-4′,4′′-
dimethoxy-5′,5′′-bis(methoxycarbonyl)-1,1-diphenyl-1-heptene (8). ADAM 8 inhibited the
cytopathic effect of HIV-1 in CEM cell culture with an EC50 value of 7.1 µM and was active
against an array of laboratory strains of HIV-1 in CEM-SS and MT-4 cells, but was inactive as
an inhibitor of HIV-2. In common with the other known non-nucleoside reverse transcriptase
inhibitors, ADAM 8 was an effective inhibitor of HIV-1 reverse transcriptase (IC50 1 µM) with
poly(rC)‚oligo(dG), but not with poly(rA)‚oligo(dT), as the template/primer. ADAM 8 was
inactive against HIV-1 reverse transcriptases containing non-nucleoside reverse transcriptase
inhibitor resistance mutations at residues 101, 106, 108, 139, 181, 188, and 236, while it
remained active against enzymes with mutations at residues 74, 98, 100, 103, and at 103/181.
An AZT-resistant virus having four mutations in reverse transcriptase was more sensitive to
inhibition by ADAM 8 than the wild-type HIV-1. In addition, ADAM 8 displayed synergistic
activity with AZT, but lacked synergy with ddI. ADAM 8 or a structurally related analog may
therefore be useful as an antiviral agent in combination with AZT or with other NNRTIs that
are made ineffective by mutations at residues which do not confer resistance to ADAM 8.
The non-nucleoside HIV-1 reverse transcriptase in-
hibitors (NNRTIs) constitute a large and structurally
diverse set of compounds having potential value in the
treatment of AIDS.1-5 Several of the more familiar
types of compounds in this class include [(hydroxy-
ethoxy)methyl](phenylthio)thymine (HEPT),6 tetrahy-
droimidazobenzodiazepinone (TIBO),7 dipyridodiazepi-
none (nevirapine),8 pyridinone,9 bis(heteroaryl)piperazine
(BHAP),10 (tert-butyldimethylsilyl)spiroaminooxathiole
dioxide (TSAO),11 and R-anilinophenylacetamide (R-
APA)12 derivatives. A number of other more recent
additions to the group include oxathiin carboxanilide
(OC or UNIROYAL)13,14 and (phenylethyl)thioureido-
thiazole (PETT)15 derivatives, as well as other com-
pounds.16-37 In addition, several natural products in
the calanolide38 and inophyllum39 families have been
identified as non-nucleoside HIV-1 reverse transcriptase
inhibitors, and halogenated derivatives of the natural
product gomisin J have displayed similar activity.40 All
of these substances inhibit HIV-1 reverse transcriptase
specifically through an allosteric mechanism involving
binding to a pocket close to, but separate from, the
deoxyribonucleoside triphosphate binding site.4,41 Al-
though the initial excitement concerning the potential
therapeutic use of the NNRTIs in the treatment of AIDS
has waned somewhat due to the rapid emergence of
resistant viral strains following their administration,
more recent work has suggested several strategies by
which this problem might be circumvented. These
include switching to another NNRTI to which the virus
has remained sensitive, using higher doses of the
NNRTI against the resistant strain,42,43 and employing
combinations of agents which elicit mutations that
counteract one another.2,44 There is therefore a definite
need for additional types of NNRTIs which might elicit
unique patterns of resistance mutations.
During our prior work on the design and synthesis of
anti-HIV agents related to cosalane (1),45-47 we ob-
served the prevention of the cytopathic effect of HIV-1
in cell culture by an analog 848 (Scheme 1) that did not
fit the general structural pattern expected for active
compounds in this series. In the cosalane series,
antiviral activity had generally been observed for com-
pounds having free carboxylic acid groups, and the
corresponding methyl esters were inactive. Compound
8, which does inhibit the cytopathic effect of HIV-1 in
cell cultures, is obviously an exception to this rule. In
addition, cosalane (1) appears to act, at least in part,
by inhibition of gp120-CD4 binding.46 When tested for
inhibition of gp120-CD4 binding, however, compound 8
was completely inactive. It consequently seemed to act
† Purdue University.
‡ Antiviral Drug Mechanisms.
§ Virology Research Group.
X Abstract published in Advance ACS Abstracts, J uly 1, 1996.
S0022-2623(96)00082-9 CCC: $12.00 © 1996 American Chemical Society

3218 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 16
Cushman et al.
Sch em e 1
Sch em e 2
Sch em e 3
by a mechanism distinct from that of cosalane, and
studies were therefore undertaken to establish its mode
of action and to delineate some of the structural
parameters associated with its anti-HIV activity. As
disclosed in the present report, compound 8 has been
found to be a novel NNRTI.
by reaction with an appropriate Wittig reagent, and the
tert-butyldimethylsilyl group was then removed with
tetra-n-butylammonium fluoride in THF to afford com-
pound 19.50 The resulting primary alcohol 19 was then
converted to the desired bromide 20 with triphenyl-
phosphine and carbon tetrabromide in boiling aceto-
nitrile.51
The amides 21 and 23 were synthesized by conversion
of the diacid 14 to its acid chloride with oxalyl chloride
in benzene at reflux, followed by treatment with am-
monia and dimethylamine, respectively. Amide 22 was
prepared by treatment of the diester 11 with methyl-
amine in refluxing aqueous methanol.
Ch em istr y
The syntheses of various alkenyldiarylmethanes (AD-
AMs) are outlined in Schemes 1-4. Methylation of the
substituted diarylmethanes 249 and 346 with dimethyl
sulfate in the presence of potassium carbonate in
refluxing acetone afforded intermediates 4 and 5, which
were oxidized with chromium trioxide in acetic anhy-
dride to yield benzophenones 6 and 7. The brominated
ADAMs 8-10 were synthesized from 6 using the ap-
propriate Wittig reagents, while the chlorinated ADAM
11 was prepared similarly from 7. Demethylation of the
hexenyl compound 8 with trimethylsilyl chloride and
sodium iodide in refluxing acetonitrile, followed by
conversion of the resulting carboxylic acids to their
ammonium salts, afforded 12. Intermediate 11 was
demethylated with boron tribromide in methylene chlo-
ride, and the resulting carboxylic acids were converted
to their ammonium salts to afford 13. The diacid 14
was obtained by selective demethylation of the two ester
functionalities of 11 with potassium hydroxide in re-
fluxing methanol.
An analog 17, containing two hydroxamic acid groups,
was prepared as outlined in Scheme 2. Esterification
of the two carboxylic acid groups of 15 with methanol
under acidic conditions gave the diester 16. Treatment
of intermediate 16 with hydroxylamine in aqueous
dioxane then yielded the desired compound 17.
The synthesis of ADAM 20, containing a brominated
side chain, is detailed in Scheme 3. The tert-butyldi-
methylsilylated intermediate 18 was prepared from 7
In order to investigate the effect of changing the
substitution pattern of the aromatic rings on the
antiviral activity, compound 28 (Scheme 4) was pre-
pared in which the two methoxyl groups present in 8
have been moved to positions ortho to the carbon atom
linking the two aromatic rings. Treatment of 5-bromo-
salicylic acid (24) with formaldehyde under acidic
conditions yielded the substituted diphenylmethane 25,
which was methylated with dimethyl sulfate and potas-

Alkenyldiarylmethanes as Anti-HIV-1 Agents
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 16 3219
Ta ble 2. Range of Activity of Compound 8 against HIV-1
Isolates in Cell Culture-Based Assays
Sch em e 4
cells
virus
EC50 (µM)a
IC50 (µM)
138 ( 12
CEM-SS
CEM-SS
MT-4
MT-4
MT-4
MT-4
MT-4
Mo/Ma
Mo/Ma
PBL
HIV-1RF
HIV-2
HIV-1IIIB
9.2 ( 2.8
Inactive
16 ( 4.3
14 ( 0.5
151
6.0 ( 3.4
0.56 ( 0.39
5.5 ( 3.5
1.85 ( 0.9
6.6 ( 4.8
136
>200
>200
>200
>200
>200
>200
>200
93 ( 25
>200
b
HIV-1A17
HIV-1N119
HIV-16R
HIV-16S
HIV-1Ba-L
HIV-1ADA
HIV-1WEJ O
SIV
c
MT-4
a
EC50 values were determined in an XTT-based cytopathicity
assay. Pyridinone-resistant. c Nevirapine-resistant.
b
to be 8. The EC50 value of 7.1 ( 5.0 µM represents an
average of seven determinations, and the range of
values was from 2.5 to 18 µM.
Replacement of the two bromine atoms in 8 by the
two chlorines in 11 resulted in a decrease in the anti-
HIV-1 potency (EC50 16.0 ( 4 µM). Substitution of the
hexenyl chain of 11 with an ω-bromopropenyl side chain
in 20 was accompanied by an increase in the anti-HIV-1
potency (EC50 8.0 ( 0.2 µM), but there was also a
significant increase in the cytotoxicity (IC50 18 ( 3 µM).
The increase in cytotoxicity seen with 20 may result
from its potential ability to nonspecifically alkylate
nucleophilic biological macromolecules through displace-
ment reactions involving the primary bromide.
Various other modifications of 8 were tried, all of them
resulting in inactive compounds. Lengthening the
alkenyl chain from C-6 in 8 to C-10 in 9 and C-16 in 10
resulted in loss of activity. Removal of the four O-
methyl groups of 8 and 11, resulting in 12 and 13, was
accompanied by a complete loss of activity. Likewise,
removal of the two ester methyl groups from 11,
resulting in 14, also caused total loss of anti-HIV-1
activity. These results emphasize the importance of the
O-methyl groups for anti-HIV-1 activity. The dihydrox-
amic acid derivative 17 was also inactive. Replacement
of the two methyl ester groups of 8 with two carbamoyl,
N-methylcarbamoyl, or N,N-dimethylcarbamoyl groups
results in the amides 21, 22, and 23, all of which were
inactive. Moving the two methoxyls from the para
positions relative to the bridging carbon in 8 to the ortho
positions in 28 was also associated with a loss of anti-
HIV-1 activity, indicating the importance of the substi-
tution patterns of the two aromatic rings. It therefore
appears that the anti-HIV activity of 8 is associated with
a relatively high degree of structural specificity, since
a variety of structural changes resulted in complete loss
of anti-HIV-1 activity.
The range of activity of ADAM 8 was tested against
a variety of laboratory strains of HIV-1 in CEM-SS and
MT4 cells. As shown in Table 2, ADAM 8 was active
against a wide range of HIV-1 strains, including one
(HIV-1A17) which is resistant to pyridinone non-nucleo-
side reverse transcriptase inhibitors. However, ADAM
8 was much less active against a nevirapine-resistant
strain of HIV-1 (HIV-1N119) as well as simian immuno-
deficiency virus (SIV) in MT-4 cells. The inactivity of
ADAM 8 against HIV-2 is consistent with its classifica-
tion as an HIV-1-specific non-nucleoside reverse tran-
scriptase inhibitor.
Ta ble 1. Anti-HIV-1 Activities of 1,1-Diphenyl-1-alkenes
a
b
compd
EC50 (µM)
IC50 (µM)
IC50/EC50
15
8
9
7.1 ( 5.0
>30
110 ( 22
>30
10
11
12
13
14
17
20
21
22
23
28
>28
>28
16.0 ( 4.4
>29.1
150 ( 10
>91.5
84 ( 10
37 ( 2
18 ( 3
31 ( 3
21 ( 0.2
21 ( 0.2
44 ( 4
>1.8
NAc
>91.5
NAc
NAc
8.0 ( 0.2
2.2
NAc
NAc
NAc
NAc
a
The EC50 is the 50% inhibitory concentration for cytopathicity
of HIV-1RF in CEM-SS cells. The values are averages of at least
b
two determinations. The IC50 is the 50% cytotoxic concentration
for mock-infected CEM cells. The values are the averages of at
least two determinations. c No activity shown at concentrations
below the IC50 value.
sium carbonate to afford 26. Oxidation of 26 with
chromium trioxide in acetic anhydride gave the ben-
zophenone derivative 27, which was then converted to
the desired product 28 using the appropriate Wittig
reagent.
Biologica l Resu lts a n d Discu ssion
The ADAMs 8-14, 17, 20-23, and 28 were tested for
prevention of the cytopathic effect of HIV-1 and for
cytotoxicity in CEM-SS cells, and the results are listed
in Table 1. The most potent ADAM, as well as the one
with the highest “therapeutic index” (IC50/EC50), proved
ADAM 8 was tested for anti-HIV activity in combina-
tion with AZT (3′-azido-3′-deoxythymidine) using the in

3220 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 16
Cushman et al.
scriptase. With poly(rC)‚oligo(dG) as the template/
primer, ADAM 8 inhibited HIV-1 reverse transcriptase
with an IC50 value of 0.38 ( 0.04 µM (mean ( 1 SD, n
) 3). However, it was inactive as an inhibitor of HIV-1
reverse transcriptase with poly(rA)‚oligo(dT) as the
template/primer. The greater sensitivity to inhibition
with poly(rC)‚oligo(dG) as the template/primer is char-
acteristic of the HIV-1-specific non-nucleoside reverse
transcriptase inhibitors.7,9,52-55
Although the non-nucleoside HIV-1 reverse tran-
scriptase inhibitors constitute a large and structurally
diverse set of compounds, there are nevertheless certain
generalities that can be made with regard to their
structures. Many of them possess two aromatic ring
systems that have heteroatoms or functional groups
attached to them that can accept hydrogen bonds. In a
number of them, the two aromatic systems are attached
to the same atom. Examples include the well-known
inhibitor nevirapine (29),8 as well as benzophenone 30,33
isoindolone 31,19 benzylpyrrole 32,27 diaryl sulfone 33,23
diaryl ketone 34,31 and related compounds. In the
HEPT 356 and DABO 3656 derivatives, one of the
aromatic rings is replaced by a pyrimidinedione ring.
The ADAM derivative 8 also has two aromatic rings
attached to one atom (carbon), and it therefore bears
some resemblance to the known NNRTIs 29-36.
F igu r e 1. Combination antiviral activity of ADAM 8 with
AZT in CEM-SS cells infected with HIV-1 (IIIB): the three-
dimensional synergy plot demonstrating the synergistic activ-
ity of the two compounds.
F igu r e 2. Combination antiviral activity of ADAM 8 with ddI
in CEM-SS cells infected with HIV-1 (IIIB): the three-
dimensional synergy plot demonstrating the lack of synergistic
activity of the two compounds.
vitro XTT anti-HIV assay. Eight concentrations of
ADAM 8 were tested in all combinations with five
concentrations of AZT. Effects of the drug combination
were calculated on the basis of the activity of the two
compounds when tested alone. The results of these
assays demonstrated that the combined antiviral activ-
ity was greater than that predicted for additivity (Figure
1). The peak above the plane represented a maximal
antiviral activity (protection from HIV-induced killing)
from a combination of ADAM 8 and AZT which was
nearly 50% greater than would have been expected if
the antiviral effects were merely additive. No antago-
nism was detected at any concentration tested.
A
similar study involving ADAM 8 and ddI demonstrated
a lack of synergistic activity (Figure 2).
Insight into the process of nevirapine (29) binding to
HIV-1 reverse transcriptase has been gained through
comparison of the crystal structure of the complex of
nevirapine with HIV-1 reverse transcriptase41 and the
crystal structure of the unliganded enzyme.57 The
results of these studies show that a hydrophobic binding
pocket is created in the p66 palm domain of HIV-1
reverse transcriptase during the binding of the inhibitor.
This involves the withdrawal of Trp-229 from the
In order to determine the mechanism of action of
ADAM 8, it was tested in a number of assays represen-
tative of important events in the replication cycle of
HIV-1. These included virus attachment to the cell,
gp120-CD4 binding, syncytium formation, reverse tran-
scription, integration, and protease. The results, which
are detailed in Table 3, indicate that the mechanism of
action of ADAM 8 is inhibition of HIV-1 reverse tran-

Alkenyldiarylmethanes as Anti-HIV-1 Agents
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 16 3221
Ta ble 3. Mechanism Studies of the Anti-HIV-1 Activity of
ADAM 8
Ta ble 5. ADAM 8 Activity against HIV-1 Viruses Containing
NNRTI Resistance Mutations Obtained by in Vitro Biological
Selection (EC50 Values, µM)
a
parameter
virion attachment
syncytia formation
gp 120-CD4 interaction
RT enzymatic activity (rAdT)
(rCdG)
IC50 (µM)
enzyme
ADAM 8 (EC50
)
AZT (EC50
)
fold resistancea
b
NI100
NI25
NI100
NI100
0.38
IIIBb
15.5
5.29
0.01
-
OC/100c
0.005
0.008
0.004
0.03
0.02
0.01
0.01
0.002
0.01
Sd
Thiazol/108e
TIBO/108f
Calo/139g
DPS/181h
UC38/181c
HEPT/181i
A17/103/181j
HEPT/236
Cos/Rk
>100
>100
>100
>100
>100
85.9
>6
>6
>6
>6
>6
5.5
S
Integrase
>200
protease enzymatic activity (IC50
)
37
p7 nucleocapsid protein zinc fingers (RFU%)c
cell-to-cell transmission
NI25
NI
17.9
a
Mechanistic studies were quantitated as described in Materi-
als and Methods. IC50 values (drug concentration providing 50%
inhibition of the indicated activity) were derived from graphs in
which each point represented the mean of at least three replicates.
>100
>6
S
11.2
0.005
a
Fold resistance (the EC50 of ADAM 8 against the mutant virus
divided by the EC50 of ADAM 8 against the wild-type virus)
indicates the level of loss of sensitivity of the mutant virus to the
b
NI indicates that no inhibition of activity was observed at the
indicated high test concentration. c The RFU% value represents
the percentage decrease in the relative fluorescence intensity of
the zinc finger after treatment of the p7 nucleocapsid protein with
25 µΜ ADAM 8 for 10 min.
b
antiviral action of ADAM 8. Wild-type enzyme. c Oxathiin car-
d
boxanilide resistant. Sensitive (no resistance). e Thiazolobenz-
imidazole resistant. f Tetrahydroimidazobenzodiazepinone resis-
g
h
tant. Calanolide resistant. Diphenyl sulfonate resistant.
i
j
Ta ble 4. ADAM 8 Activity against HIV-1 Viruses Containing
NNRTI Resistance Mutations Obtained by Site-Directed
Mutagenesis (EC50 Values, µM)
(Hydroxyethoxy)methyl](phenylthio)thymine resistant. Pyridi-
none resistant. Cosalane resistant.
k
sistance mutations are located close to the bound
inhibitor,60 and it therefore seems reasonable to assume
that the NNRTI resistance mutations conferring resis-
tance to ADAM 8 may also be located close to bound
ADAM 8. These amino acid residues are located at
positions 101, 106, 108, 139, 179, 181, 188, and 236.
There are also several known NNRTI resistance
mutations located in the NNRTI binding pocket that do
not confer significant resistance to ADAM 8. These
include residues 98, 100, and 103. The enzyme with
two mutations at residues 103 and 181 also remains
sensitive to inhibition by ADAM 8. It is generally
recognized that NNRTI resistance mutations do not
confer resistance to all NNRTIs, and that a potentially
viable strategy for dealing with NNRTI resistance may
be to simply switch to another NNRTI that remains
effective against the resistant virus.2,44 The A98G
mutation is known to confer resistance to pyridinone
L-697,66161 and nevirapine,62 while the OC/100 virus
is oxathiin carboxanilide resistant. The A17/103/181
virus is resistant to nevirapine. It is of interest that
the virus containing the 4xAZT enzyme, which has four
mutations in its reverse transcriptase and is resistant
to AZT, displayed increased sensitivity to ADAM 8
relative to the wild-type virus containing the NL4-3
enzyme.
Figure 3, which was obtained by erasing nevirapine
from the structure of the nevirapine-RT complex, shows
the amino acid residues in the p66 subunit surrounding
the NNRTI binding pocket.41 Mutation of the red- and
yellow-colored residues in Figure 3 results in resistance
to certain of the known NNRTIs. Mutation of the
yellow-colored residues in Figure 3 also results in
resistance to ADAM 8, while mutation of the red-colored
residues does not confer resistance to ADAM 8 (Table
4 and 5). The yellow-colored lysine residue 101 is
located at the assumed entrance to the binding pocket,
near the red-colored lysine residue 103. The two yellow-
colored tyrosine residues are Tyr 181 and Tyr 188, while
the three yellow valine residues are Val 106, Val 108,
and Val 179. The yellow-colored proline residue is Pro
236. The red-colored alanine residue is Ala 98, and the
red-colored leucine residue is Leu 100. The mutation
at residue 139 conferring resistance to ADAM 8 is
located on the p51 subunit and is not shown. Taken
enzyme
ADAM 8 (EC50
)
AZT (EC50)
fold resistanceb
NL4-3a
L74V
A98G
K101E
K103N
V106A
V179D
Y181C
Y188C
4xAZTd
12.3
5.47
24.1
0.05
0.03
0.03
1
0.01
0.01
0.02
0.003
0.05
0.98
-
Sc
S
>100
>8
S
23.6
>100
>100
>100
>100
0.93
>8
>8
>8
>8
vS
a
b
Wild-type enzyme. Fold resistance (the EC50 of ADAM 8
against the mutant virus divided by the EC50 of ADAM 8 against
the wild-type virus) indicates the level of loss of sensitivity of the
mutant virus to the antiviral action of ADAM 8. c Sensitive (no
d
resistance). AZT resistant, with four mutations.
binding site as well as conformational changes of the
Tyr-181 and Tyr-188 side chains. Inhibitor binding may
be driven by π-π interactions between the ligand and
aromatic amino acid residues present at the binding
site.4 The filling of the hydrophobic binding pocket by
the inhibitor is associated with domain shifts in the
enzyme that may alter oligonucleotide binding. It is
possible that similar processes occur during the binding
of ADAM 8, as well as the related compounds 30-36, to
HIV-1 reverse transcriptase. The orientations of the
two aromatic systems in these compounds fit a “but-
terfly-like” model recently proposed on the basis of a
comparison of the structures of several non-nucleoside
HIV-1 reverse transcriptase inhibitors.58 In the case of
ADAM 8, the hexenyl side chain points toward a
“lipophilic site” proposed by the model.
In order to identify the amino acid residues of HIV-1
reverse transcriptase that may be involved in ADAM
binding, ADAM 8 was tested for inhibitory activity
against a variety of HIV-1 strains containing NNRTI
resistance mutations in the reverse transcriptase en-
zyme. The viruses were obtained either by site-directed
mutagenesis or by in vitro biological selection, and the
results of these studies are listed in Tables 4 and 5.
Comparison of these data with the published structures
of the NNRTI binding pocket shows that the mutations
conferring resistance to ADAM 8 are clustered within
the amino acid residues comprising the pocket (see
Figure 3).4,41,59,60 In the case of R-APA, it has been
documented that the commonly identified NNRTI re-

3222 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 16
Cushman et al.
F igu r e 3. The non-nucleoside HIV-1 reverse transcriptase inhibitor (NNRTI) binding site. Mutation of the yellow- and red-
colored residues results in resistance to known NNRTIs. Mutation of the yellow residues also results in resistance to ADAM 8,
while mutation of the red residues does not confer resistance to ADAM 8.
3,3′-Dich lor o-4,4′-d im eth oxy-5,5′-bis(m eth oxyca r bon -
yl)d ip h en ylm eth a n e (5). This intermediate was prepared
from 3 as previously described.46
together, the cluster of yellow amino acid residues
depicted in Figure 3 indicate the geometrical relation-
ships of the residues that are likely to be involved in
ADAM 8 binding, and they circumscribe a well-defined
cavity which most likely constitutes the ADAM 8
binding site.
Additional studies will be required in order to further
define the structural parameters associated with the
HIV-1 reverse transcriptase inhibitory activity of the
compounds in the ADAM series, as well as to maximize
the anti-HIV potency and therapeutic index. It will also
be of interest to define the pattern of resistance muta-
tions associated with the use of ADAM 8.
The fact that ADAM 8 displays synergistic anti-HIV-1
activity with AZT and is actually more effective against
an AZT-resistant virus suggests that ADAM 8 or a
closely related analog may prove to be useful in treating
HIV-1 infections in combination with AZT. It may also
find some utility against viral strains that have been
made resistant to other NNRTIs by mutations that do
not confer resistance to ADAM 8.
3,3′-Dibr om o-4,4′-d im eth oxy-5,5′-bis(m eth oxyca r bon -
yl)d ip h en yl Keton e (6). Diphenylmethane 4 (6.02 g, 12
mmol) was partially dissolved in acetic anhydride (110 mL),
the solution was cooled in an ice bath, and chromium trioxide
(5.05 g) was added in small portions with stirring. The
mixture was stirred at ambient temperature for 3 h and then
for 2 h at reflux and was filtered after cooling. The chromium
salts and ketone 6 were filtered off, washed with chloroform,
and then extracted twice by stirring the solid with chloroform
(80 mL) and refiltering the suspension. The solvents were
removed in vacuo from the combined chloroform washings and
extracts, and the residue was crystallized from a chloroform-
hexane mixture to yield the product 6 (4.84 g 78%): mp 130-
131 °C; IR (KBr) 2954, 1742, 1657, 1589, 1472, 1438, 1318,
1268, 1201, 1090, 986 cm-1; 1H NMR (CDCl3, 200 MHz) δ 8.16
(d, J ) 2.2 Hz, 2 H), 8.12 (d, J ) 2.2 Hz, 2 H), 4.03 (s, 6 H),
3.95 (s, 6 H); CIMS m/ e (relative intensity) 517 (MH+, 100),
485 (25). Anal. (C19H16Br2O7) C, H.
3,3′-Dich lor o-4,4′-d im eth oxy-5,5′-bis(m eth oxyca r bon -
yl)d ip h en yl Ket on e (7). This intermediate was prepared
from 5 as previously described.46
3′,3′′-Dibr om o-4′,4′′-d im eth oxy-5′,5′′-bis(m eth oxyca r bo-
n yl)-1,1-d ip h en yl-1-h ep ten e (8). n-Hexyltriphenylphospho-
nium bromide63 (1.43 g, 3.35 mmol) was suspended in dry THF
(25 mL), the mixture was cooled in an ice bath, and sodium
bis(trimethylsilyl)amide (1 M solution in THF, 3.35 mL) was
added dropwise. The mixture was stirred for 20 min, and a
solution of the bromo ketone 6 (1.554 g, 3.01 mmol) in THF
(12 mL) was added dropwise. The ice bath was removed, and
the reaction mixture was stirred 1 h at 60 °C and 20 h at
ambient temperature. The reaction was quenched with am-
monium chloride solution, the THF phase was separated and
the aqueous phase extracted with ether. The combined organic
extracts were dried (sodium sulfate) and evaporated to dry-
ness. Flash chromatography on silica gel (hexane-ethyl
acetate, 4:1) yielded solid that was recrystallized from ethanol
to afford a crystalline product (1.593 g, 91%): mp 52-53 °C;
IR (KBr) 2946, 2860, 1732, 1472, 1435, 1288, 1250, 1209, 1086,
Exp er im en ta l Section
Gen er a l. Melting points were determined in capillary
tubes on a Mel-Temp apparatus and are uncorrected. Spectra
were obtained as follows: CI mass spectra on a Finnegan 4000
spectrometer; FAB mass spectra and EI mass spectra on a
Kratos MS50 spectrometer; 1H NMR spectra on Varian VXR-
500S and XL-200A spectrometers; IR spectra on a Beckman
IR-33 spectrometer or on a Perkin-Elmer 1600 series FTIR.
Microanalyses were performed at the Purdue Microanalysis
Laboratory, and all values were within (0.4% of the calculated
compositions.
3,3′-Dibr om o-4,4′-d im eth oxy-5,5′-bis(m eth oxyca r bon -
yl)d ip h en ylm eth a n e (4). 3,3′-Dibromo-5,5′-dicarboxy-4,4′-
dihydroxydiphenylmethane (2)49 (8.179 g, 18.34 mmol) was
partially dissolved in acetone (250 mL), potassium carbonate
(20.09 g) was added followed by dimethyl sulfate (9.5 mL), and
the mixture was heated at reflux with stirring for 24 h.
Acetone was removed in vacuo, water (100 mL) was added,
and the mixture was extracted with methylene chloride (6 ×
30 mL). The combined organic extracts were washed with 2%
potassium hydroxide (50 mL), brine, dried (sodium sulfate),
and evaporated to dryness. The residue was crystallized from
a chloroform-hexane mixture to afford product 4 (7.75 g,
84%): mp 107-108 °C; IR (KBr) 2948, 1725, 1595, 1553, 1472,
1433, 1314, 1251, 1202, 1089, 996, 926, 876, 827, 800, 717
1
997, 726 cm-1; H NMR (CDCl3, 200 MHz) δ 7.53 (d, J ) 2.2
Hz, 2 H), 7.495 (d, J ) 2.2 Hz, 1 H), 7.48 (d, J ) 2.2 Hz, 1 H),
6.06 (t, J ) 7.6 Hz, 1 H), 3.99 (s, 3 H), 3.92 (s, 6 H), 3.92 (s, 3
H), 2.08 (m, 2 H), 1.45 (m, 2 H), 1.26 (m, 4 H), 0.88 (t, J ) 6.7
Hz, 3 H); CIMS m/ e (relative intensity) 585 (MH+, 85), 553
(100). Anal. (C25H28Br2O6) C, H.
3′,3′′-Dibr om o-4′,4′′-d im eth oxy-5′,5′′-bis(m eth oxyca r bo-
n yl)-1,1-d ip h en yl-1-u n d ecen e (9). n-Decyltriphenylphos-
phonium bromide64 (1.15 g, 2.62 mmol) was suspended in dry
THF (22 mL), the mixture was cooled in an ice bath, and
sodium bis(trimethylsilyl)amide (1 M solution in THF, 2.62
mL) was added dropwise. The mixture was stirred for 20 min,
and a solution of ketone 6 (1.082 g, 2.096 mmol) in THF (18
1
cm-1; H NMR (CDCl3, 200 MHz) δ 7.55 (d, J ) 2.2 Hz, 2 H),
7.50 (d, J ) 2.2 Hz, 2 H), 3.92 (s, 6 H), 3.91 (s, 6 H), 3.88 (s, 2
H); CIMS m/ e (relative intensity) 503 (MH+, 76), 471 (100),
257 (63). Anal. (C19H18Br2O6) C, H.

Alkenyldiarylmethanes as Anti-HIV-1 Agents
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 16 3223
mL) was added dropwise. The ice bath was removed, and the
reaction mixture was stirred 24 h at ambient temperature.
The reaction was quenched with ammonium chloride solution,
the THF phase was separated, and the aqueous phase was
extracted with ether. The combined organic extracts were
dried (sodium sulfate) and evaporated to dryness, and the
residue was purified by flash chromatography on silica gel
(hexane-ethyl acetate, 4:1) and crystallization from ethanol
to yield the product 9 (1.25 g, 93%): mp 69-70 °C; IR (KBr)
2928, 2855, 1734, 1471, 1435, 1364, 1288, 1251, 1208, 1087,
extracts were washed with brine and dried (sodium sulfate)
to yield a foam (191 mg, 68%), which was crystallized from
ethyl acetate: mp 159-160 °C; 1H NMR (DMSO-d6, 200 MHz)
δ 7.44 (d, J ) 2.0 Hz, 1 H), 7.36 (d, J ) 1.9 Hz, 1 H), 7.18 (d,
J ) 2.0 Hz, 1 H), 7.10 (d, J ) 1.9 Hz, 1 H), 5.75 (t, J ) 7.4 Hz,
1 H), 2.02 (m, 2 H), 1.37 (m, 2 H), 1.22 (m, 4 H), 0.82 (t, J )
6.1 Hz, 3 H); FABMS m/ e (relative intensity) 529 (MH+, 2).
Anal. (C21H20Br2O6‚2H2O) C, H. The diammonium salt 12 was
prepared by dissolving the diacid in ammonium hydroxide and
then evaporating the water and excess ammonia.
998, 727 cm-1
;
1H NMR (CDCl3, 200 MHz) δ 7.53 (m, 2 H),
Dia m m on iu m 3′,3′′-Dich lor o-5′,5′′-d ica r boxy-4′,4′′-d i-
h yd r oxy-1,1-d ip h en yl-1-h ep ten e (13). A solution of meth-
oxy ester 11 (148.6 mg, 0.3 mmol) in dry methylene chloride
(2.5 mL) was added via septum under an argon atmosphere
to a stirred solution of boron tribromide (1 M) in methylene
chloride (1.6 mL) cooled in a dry ice-acetone bath. The cooling
bath was removed after 2 h, and stirring continued at ambient
temperature for 2 days. More BBr3 (0.8 mL) was added on
the second day. The reaction was quenched with water (2 mL),
stirring was continued for 30 min, and the product was
extracted with 20% aqueous KOH. The alkaline solution was
acidified on cooling with concentrated hydrochloric acid and
the product extracted with ethyl acetate. The organic extracts
were washed with brine, dried (sodium sulfate), and concen-
trated in vacuo. The product (19 mg, 32%) was crystallized
from methylene chloride: mp 234-236 °C; IR (KBr) 3500-
2500, 2926, 2856, 1670, 1600, 1443, 1232, 1179, 901, 799, 715
cm-1; 1H NMR (CDCl3, 200 MHz) δ 7.68 (m, 2 H), 7.50 (d, J )
1.6 Hz, 1 H), 7.41 (d, J ) 1.6 Hz, 1 H), 6.13 (t, J ) 7.2 Hz, 1
H), 2.11 (m, 2 H), 1.48 (m, 2 H), 1.28 (m, 14 H), 0.85 (t, J )
6.3 Hz, 3 H); FABMS m/ e (relative intensity) 438 (M+, 2), 307
(10). Anal. (C21H20Cl2O6‚1/2H2O) C, H. The ammonium salt
13, mp 150 °C, was prepared by dissolving the diacid in
ammonium hydroxide and evaporating the water and excess
ammonia.
7.49 (m, 2 H), 6.06 (t, J ) 7.5 Hz, 1 H), 3.99 (s, 3 H), 3.92 (s,
9 H), 2.08 (m, 2 H), 1.44 (m, 2 H), 1.25 (m, 12 H), 0.88 (t, J )
5.3 Hz, 3 H); CIMS m/ e (relative intensity) 641 (MH+, 80),
609 (100). Anal. (C29H36Br2O6) C, H.
3′,3′′-Dibr om o-4′,4′′-d im eth oxy-5′,5′′-bis(m eth oxyca r bo-
n yl)-1,1-d ip h en yl-1-h ep t a d ecen e (10). n-Hexadecyltri-
phenylphosphonium bromide65 (1.706 g, 3.006 mmol) was
dissolved in dry THF (32 mL), the solution was cooled in an
ice bath, and sodium bis(trimethylsilyl)amide (1 M solution
in THF, 3 mL) was added dropwise. The mixture was stirred
for 20 min, and a solution of ketone 6 (1.393 g, 2.7 mmol) in
THF (12 mL) was added dropwise. The ice bath was removed,
and the reaction mixture was stirred for 28 h at ambient
temperature. The reaction was quenched with ammonium
chloride solution, the THF phase was separated, and the
aqueous phase was extracted with ether. The combined
organic extracts were dried (sodium sulfate) and evaporated
to dryness. Flash chromatography on silica gel (hexane-ethyl
acetate, 6:1) afforded an oil, which crystallized from ethanol
to yield the product 10 (1.15 g, 59%): mp 43 °C (ethanol); IR
(KBr) 2923, 2852, 1734, 1590, 1470, 1435, 1288, 1251, 1209,
1
1087, 999, 727 cm-1; H NMR (CDCl3, 200 MHz) δ 7.52 (d, J
) 2.3 Hz, 2 H), 7.49 (d, J ) 2.3 Hz, 1 H), 7.48 (d, J ) 2.3 Hz,
1 H), 6.86 (t, J ) 7.4 Hz, 1 H), 3.99 (s, 3 H), 3.92 (s, 9 H), 2.08
(m, 2 H), 1.42 (m, 2 H), 1.25 (m, 24 H), 0.875 (t, J ) 6.5 Hz, 3
H); CIMS m/ e (relative intensity) 725 (MH+ - MeOH, 100).
Anal. (C35H48Br2O6) C, H.
3′,3′′-Dich lor o-5′,5′′-d ica r b oxy-4′,4′′-d im e t h oxy-1,1-
d ip h en yl-1-h ep ten e (14). Diester 11 (35 mg, 0.071 mmol)
was heated for 2 h under reflux in a solution of potassium
hydroxide (28 mg, 0.5 mmol) in methanol (2 mL). Methanol
was evaporated in vacuo, water was added, and the solution
was acidified with hydrochloric acid (1 M, 1 mL). The white
precipitate was extracted with chloroform (5 × 2 mL), and the
combined extracts were washed with brine, dried (sodium
sulfate), and evaporated in vacuo to yield a white solid (31
mg, 100%), which was crystallized from methylene chloride:
mp 204 °C; IR (KBr) 3425, 3500-2500, 1696, 1597, 1555, 1296
cm-1; 1H NMR (200 MHz, acetone-d6) δ 7.58 (d, J ) 2.1 Hz, 1
H), 7.58 (d, J ) 2.1 Hz, 1 H), 7.49 (d, J ) 2.1 Hz, 2 H), 6.28 (t,
J ) 7.5 Hz, 1 H), 3.96 (s, 3 H), 3.90 (s, 3 H), 2.12 (m, 2 H), 1.5
(m, 2 H), 1.28 (m, 4 H), 0.85 (t, J ) 6.6 Hz, 3 H); CIMS m/ e
3′,3′′-Dich lor o-4′,4′′-d im eth oxy-5′,5′′-bis(m eth oxyca r bo-
n yl)-1,1-d ip h en yl-1-h ep ten e (11). Sodium hydride (54 mg,
1.35 mol, 60% dispersion in mineral oil) was washed with
n-hexane (3 × 5 mL). Dimethyl sulfoxide (2 mL) was intro-
duced via a syringe, and the mixture was heated at 75 °C until
the evolution of hydrogen ceased. The clear solution was
cooled in an ice-water bath, and a solution of the n-hexyl-
triphenylphosphonium bromide63 (576 mg, 1.35 mmol) in
DMSO (3 mL) was added dropwise. The resulting solution was
stirred at room temperature for 15 min. A solution of the
ketone 7 (0.86 mmol, 369 mg) in warm DMSO (6 mL) was
added dropwise and the reaction mixture heated at 55 °C for
27 h. It was then cooled in an ice bath. A solution of
ammonium chloride (143 mg) in water (5 mL) was added, and
the reaction mixture was extracted with ethyl ether (5 × 5
mL). The organic extracts were washed twice with brine, dried
over sodium sulfate, and evaporated in vacuo. Flash chroma-
tography on silica gel (230-400 mesh), eluting with n-hexane-
ethyl acetate, 4:1, yielded starting ketone (39 mg) and heptene
8 (320 mg, 75%): mp 88 °C; IR (KBr) 3046, 1732, 1595, 1364,
(relative intensity) 467 (MH+, 42), 449 (100). Anal. (C23H24
Cl2O6) C, H.
-
3′,3′′-Dich lor o-4′,4′′-d ih yd r oxy-5′,5′′-bis(m eth oxyca r bo-
n yl)-1,1-d ip h en yl-1-h ep ten e (16). The diacid 15 (98 mg,
0.11 mmol), obtained by acidification of 12 with hydrochloric
acid, was added to a solution of concentrated sulfuric acid (5
drops) in methanol (4 mL), and the mixture was heated at 90
°C for 5 days. The reaction mixture was cooled in an ice bath,
water was added, and the product was extracted with meth-
ylene chloride (3 × 3 mL). The combined extracts were washed
with 5% sodium bicarbonate solution and brine and dried
(sodium sulfate). The solvent was removed in vacuo, and the
residue was flash chromatographed on silica gel (4 g, hexane-
ethyl acetate, 4:1) to yield the ester as an oil which crystallized
from a mixture of methanol and ether to afford the product
16 (78 mg, 75%): mp 118 °C; IR (KBr) 3422, 3125, 2952, 2927,
1
1250, 1211 cm-1; H NMR (CDCl3, 500 MHz) δ 7.49 (d, J )
2.4 Hz, 1 H), 7.48 (d, J ) 2.2 Hz, 1 H), 7.32 (d, J ) 2.2 Hz, 1
H), 7.30 (d, J ) 2.4 Hz, 1 H), 6.07 (t, J ) 7.6 Hz, 1 H), 4.00 (s,
3 H), 3.93 (s, 3 H), 3.92 (s, 3 H), 3.91 (s, 3 H), 2.08 (m, 2 H),
1.44 (m, 2 H), 1.27 (m, 4 H), 0.88 (t, J ) 6.9 Hz, 3 H); CIMS
m/ e (relative intensity) 495 (MH+, 100), 463 (12). Anal.
(C25H28Cl2O6) C, H.
Dia m m on iu m 3′,3′′-Dib r om o-5′,5′′-d ica r b oxy-4′,4′′-d i-
h yd r oxy-1,1-d ip h en yl-1-h ep ten e (12). Methoxy ester 8
(0.289 g, 0.495 mmol) and sodium iodide (0.593 g) were stirred
in dry acetonitrile under argon atmosphere. Trimethylsilyl
chloride (1.27 mL, 9.9 mmol, 20 equiv) was added dropwise at
room temperature, and the reaction mixture was stirred under
reflux for 48 h. The reaction mixture was quenched with water
(3 mL) and extracted with ethyl acetate (3 × 3 mL). The
combined extracts were washed with brine, 10% sodium
sulfite, and 5% sodium bicarbonate (3 × 4 mL). The basic
extracts were acidified with concentrated hydrochloric acid,
and the product was extracted with ethyl acetate. The organic
2858, 1679, 1604, 1442, 1334, 1237, 1200, 1168, 797, 750 cm-1
;
1H NMR (CDCl3, 200 MHz) δ 11.37 (s, 1 H), 11.28 (s, 1 H),
7.53 (d, J ) 2.1 Hz, 1 H), 7.50 (d, J ) 2.2 Hz, 1 H), 7.38 (dd,
J ) 2.2, 0.3 Hz, 1 H), 7.33 (bd, J ) 2.1 Hz, 1 H), 5.96 (t, J )
7.5 Hz, 1 H), 3.93 (s, 3 H), 3.92 (s, 3 H), 2.03 (m, 2 H), 1.43 (m,
2 H), 1.23 (m, 6 H), 0.85 (t, J ) 5.1 Hz, 3 H); CIMS m/ e
(relative intensity) 467 (MH+, 1), 465 (MH+ - 2H, 3), 433 (3),
281 (100). Anal. (C23H24Cl2O6) C, H.
3′,3′′-Dich lor o-5′,5′′-bis(h yd r oxyca r ba m oyl)-4′,4′′-d ih y-
d r oxy-1,1-d ip h en yl-1-h ep ten e (17). A solution of 3 M
sodium hydroxide (3.07 mL) was added to a solution of

3224 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 16
Cushman et al.
hydroxylamine hydrochloride (275 mg, 3.95 mmol) in water
(2.8 mL), followed by a solution of the dimethyl ester 16 (74
mg, 0.158 mmol) in dioxane (2 mL). The cloudy mixture was
stirred at room temperature for 20 h, cooled in an ice bath,
acidified with 3 M hydrochloric acid (2 mL), and extracted with
ethyl acetate. The combined extracts were washed with brine
and dried (sodium sulfate). The solvent was removed in vacuo
and the residue crystallized on trituration with methylene
chloride to afford 17 (67 mg, 90%): mp 137-138 °C; IR (KBr)
3340, 3160, 2925, 1648, 1470, 1108, 978 cm-1; 1H NMR (CDCl3,
200 MHz) δ 11.21 (bs, 2 H), 8.65 (bs, 2 H), 7.53 (d, J ) 2.1 Hz,
1 H), 7.47 (d, J ) 2.1 Hz, 1 H), 7.45 (d, J ) 2.1 Hz, 1 H), 7.38
(d, J ) 2.1 Hz, 1 H), 6.14 (t, J ) 7.5 Hz, 1 H), 2.05 (m, 2 H),
1.49 (m, 2 H), 1.27 (m, 4 H), 0.84 (t, J ) 6.8 Hz, 3 H); FABMS
m/ e (relative intensity) 469 (MH+, 75), 436 (MH+ - NH2OH,
100). Anal. (C21H22N2Cl2O6) C, H, N.
[3-[(t er t -B u t y ld im e t h y ls ily l)o x y ]p r o p y l]t r ip h e n -
ylp h osp h on iu m Br om id e. 3-Bromo-3-(tert-butyldimethyl-
silyl)-1-propanol (2.7 g, 10.75 mmol) and triphenylphosphine
(2.83 g, 10.75 mmol) were dissolved in hot acetonitrile (3
mL) and heated under reflux for 30 h. The reaction mixture
was allowed to cool, the solvent removed in vacuo, and the
oily residue triturated with hexane until crystallization came
to completion. The white salt (4.995 g, 93%) was filtered the
next day, washed with hexane (3 × 5 mL), and dried in a
vacuum desiccator: mp 137-139 °C; 1H NMR (DMSO-d6, 200
MHz) δ 7.9-7.8 (m, 15 H), 3.68 (t, J ) 5.9 Hz, 2 H), 3.53 (m,
2 H), 1.67 (m, 2 H), 0.84 (s, 9 H), 0.02 (s, 6 H); EIMS 435
(M+ - Br).
for 2 h and cooled, the solvent was removed in vacuo, and the
residue was extracted with benzene (3 × 5 mL). The combined
extracts were filtered, and the solvent was removed in vacuo.
The residue was flash chromatographed on silica gel (20 g,
hexane-ethyl acetate, 4:1) and recrystallized from ethanol to
yield the bromide 20 (0.20 g, 82%): mp 68 °C; IR (KBr) 2949,
1730, 1476, 1437, 1297, 1254, 1214, 995 cm-1; 1H NMR (CDCl3,
200 MHz) δ 7.51 (m, 2 H), 7.36 (d, J ) 2.1 Hz, 1 H), 7.33 (d, J
) 2.3 Hz, 1 H), 6.08 (t, J ) 7.2 Hz, 1 H), 4.00 (s, 3 H), 3.93 (s,
9 H), 3.46 (t, J ) 6.6 Hz, 2 H), 2.69 (q, J ) 6.7 Hz, 2 H); CIMS
m/ e (relative intensity) 533 (MH+, 94), 501 (100). Anal.
(C22H21BrCl2O6) C, N.
3′,3′′-Dica r b a m oyl-5′,5′′-d ich lor o-4′,4′′-d im et h oxy-1,1-
d ip h en yl-1-h ep ten e (21). The diacid 14 (112 mg, 0.240
mmol) was suspended in dry benzene (2 mL), oxalyl chloride
(0.4 mL) was added, and the mixture was heated at reflux for
15 min. The solution was stirred for 2 h, and the solvent was
removed in vacuo. The acid chloride was dissolved in dry
benzene (4 mL), and sodium-dried ammonia was passed
through the stirred solution for 2 h. Water was added,
followed by chloroform, and the mixture was filtered from
contaminating oxalyl amide. The organic phase was washed
with a 5% solution of potassium hydroxide and then brine,
dried (sodium sulfate), and evaporated to dryness. The residue
was purified by preparative TLC on silica gel (ethyl acetate-
ethanol, 97:3), and the amide 21 (60 mg, 54%) was crystallized
from methylene chloride-hexane: mp 61-62 °C; IR (KBr)
3447, 3263, 2930, 2855, 1718, 1687, 1579, 1474, 1375, 1250,
1
991 cm-1; H NMR (acetone-d6, 200 MHz) δ 7.66 (d, J ) 1.9
Hz, 2 H), 7.52 (d, J ) 2 Hz, 1 H), 7.43 (d, J ) 1.9 Hz, 1 H),
6.32 (m, 1 H), 3.99 (s, 3 H), 3.98 (s, 3 H), 2.13 (m, 2 H), 1.51
(m, 2 H), 1.29 (m, 4 H), 0.87 (t, J ) 5.5 Hz, 3 H); FABMS m/ e
4-[(ter t-Bu t yld im et h ylsilyl)oxy]-3′,3′′-d ich lor o-4′,4′′-
d im et h oxy-5′,5′′-b is(m et h oxyca r b on yl)-1,1-d ip h en yl-1-
bu ten e (18). [3-[(tert-Butyldimethylsilyl)oxy]propyl]triphen-
ylphosphonium bromide (1.27 g, 2.46 mmol) was suspended
in dry DME (10 mL), and n-butyllithium (2.4 M, 1.03 mL) was
added dropwise. A solution of ketone 7 (1.05 g, 2.46 mmol) in
DME (12 mL) was added dropwise to the solution of the ylide,
and the mixture was stirred at 63 °C for 22 h. The cooled
reaction mixture was quenched with aqueous ammonium
chloride solution (250 mg in 10 mL of water). The aqueous
layer was extracted with ether (5 mL), and the combined
organic extracts were washed with brine and dried (sodium
sulfate). The solvent was evaporated in vacuo to afford an oil
(2.05 g), which was flash chromatographed on silica gel (90 g,
hexane-ethyl acetate, 4:1) to yield a colorless oil (0.617 g,
43%) which crystallized from ethanol: mp 40 °C; IR (KBr)
2946, 2861, 1735, 1596, 1475, 1436, 1252, 1210, 1096, 1001,
(relative intensity) 465 (MH+, 16), 448 (100). Anal. (C23H26
Cl2N2O4) C, H.
-
3′,3′′-Bis(m e t h ylca r b a m oyl)-5′,5′′-d ich lor o-4′,4′′-d i-
m eth oxy-1,1-d ip h en yl-1-h ep ten e (22). Diester 11 (140 mg,
0.295 mmol) was dissolved in hot methanol (1 mL), and
methylamine (1 mL, 40% aqueous solution) was added. The
reaction mixture was heated at reflux for 5 h, cooled, and
concentrated, and water (3 mL) was added. The product was
extracted with ether, the organic phase was washed with
water, 5% potassium hydroxide, and brine, dried (sodium
sulfate), and evaporated to dryness. The residue (130 mg,
89%) was crystallized from ether-hexane to afford the prod-
uct: mp 118-119 °C; IR (neat) 2930, 2863, 1642, 1556, 1475,
1
1390, 1264, 1165, 1085, 999, 734 cm-1; H NMR (CDCl3, 500
1
842, 777 cm-1; H NMR (CDCl3, 200 MHz) δ 7.50 (d, J ) 2.2
MHz) δ 7.82 (d, J ) 2.5 Hz, 1 H), 7.76 (d, J ) 2.2 Hz, 1 H),
7.67 (m, 2 H), 7.28 (d, J ) 2.2 Hz, 1 H), 7.22 (d, J ) 2.5 Hz, 1
H), 6.10 (t, J ) 7.0 Hz, 1 H), 3.95 (s, 3 H), 3.88 (s, 3 H), 3.02
(m, 6 H), 2.09 (m, 2 H), 1.42 (m, 2 H), 1.27 (m, 4 H), 0.86 (m,
3 H); FABMS m/ e (relative intensity) 493 (MH+, 100). Anal.
Calcd (C25H30Cl2N2O4) C, H.
Hz, 1 H), 7.46 (d, J ) 2.0 Hz, 1 H), 7.37 (d, J ) 2.0 Hz, 1 H),
7.27 (d, J ) 2.2 Hz, 1 H), 6.09 (t, J ) 7.5 Hz, 1 H), 3.97 (s, 3
H), 3.91 (s, 3 H), 3.89 (s, 6 H), 3.68 (t, J ) 6.4 Hz, 2 H), 2.30
(m, 2 H), 0.87 (s, 9 H), 0.025 (s, 6 H); FABMS m/ e (relative
intensity) 583 (MH+, 63), 551 (90), 525 (100). Anal. (C28H36
Cl2O7Si) C, H.
-
3′,3′′-Bis(d im et h ylca r b a m oyl)-5′,5′′-d ich lor o-4′,4′′-d i-
m eth oxy-1,1-d ip h en yl-1-h ep ten e (23). Compound 14 (128
mg, 0.274 mmol) was converted to the corresponding acid
chloride as described in the preparation of 21. The chloride
was dissolved in dry benzene (4 mL), and anhydrous dimeth-
ylamine was bubbled through the stirred solution at 5-10 °C
for 2 h. The solvent was removed in vacuo, water was added,
and the product was extracted with methylene chloride. The
combined extracts were washed with a 5% solution of potas-
sium hydroxide and brine, dried (sodium sulfate), and evapo-
rated to dryness. The residue was purified by flash chroma-
tography on alumina (hexane-ethyl acetate, 1:1) to yield the
product as a low-melting glass (90 mg, 63%): IR (neat) 2930,
3′,3′′-Dich lor o-4-h yd r oxy-4′,4′′-d im e t h oxy-5′,5′′-b is-
(m eth oxyca r bon yl)-1,1-d ip h en yl-1-bu ten e (19). The silyl
ether 18 (407 mg, 0.687 mmol) was stirred at 0 °C for 2.5 h in
a solution prepared by adding a 1 M solution of tetra-n-
butylammonium fluoride in THF (1.4 mL) to THF (4 mL). The
solvent was removed in vacuo, brine was added, and the
product was extracted with benzene. The combined extracts
were washed with brine, dried (sodium sulfate), evaporated
in vacuo, and flash chromatographed on silica gel to yield a
colorless oil (0.255 g, 78%): IR (neat) 3528, 3061, 2952, 2875,
2832, 1732, 1596, 1477, 1437, 1402, 1359, 1269, 1210, 1168,
1
1094, 999, 739 cm-1; H NMR (CDCl3, 200 MHz) δ 7.52 (d, J
) 2.5 Hz, 1 H), 7.52 (d, J ) 2.2 Hz, 1 H), 7.38 (d, J ) 2.2 Hz,
1 H), 7.32 (d, J ) 2.5 Hz, 1 H), 6.14 (t, J ) 7.5 Hz, 1 H), 4.00
(s, 3 H), 3.94 (s, 3 H), 3.92 (s, 3 H), 3.92 (s, 3 H), 3.75 (m, 2 H),
2.39 (m, 2 H); CIMS m/ e (relative intensity) 469 (MH+, 100),
451 (35), 437 (780). Anal. (C22H22Cl2O7) C, H.
2863, 1642, 1556, 1473, 1390, 1264, 1165, 1085, 999, 734 cm-1
;
1H NMR (CDCl3, 200 MHz) δ 7.24 (bs, 1 H), 7.18 (bs, 1 H),
6.98 (bs, 1 H), 6.91 (bs, 1 H), 6.02 (t, J ) 7.3 Hz, 1 H), 3.91 (s,
3 H), 3.83 (s, 3 H), 2.86 (s, 6 H), 2.84 (s, 6 H), 2.06 (m, 2 H),
1.39 (m, 2 H), 1.23 (m, 4 H), 0.845 (t, J ) 5.8 Hz, 3 H); FABMS
m/ e relative intensity 521 (MH+, 100), 476 (82), 433 (27).
Anal. (C27H34Cl2N2O4) C, H.
4-B r o m o -3′,3′′-d ic h lo r o -4′,4′′-d im e t h o x y -5′,5′′-b is -
(m eth oxyca r bon yl)-1,1-d ip h en yl-1-bu ten e (20). A solution
of alcohol 19 (215 mg, 0.458 mmol) and carbon tetrabromide
(304 mg, 0.916 mmol) in dry acetonitrile (3 mL) was heated
under reflux with stirring, and a solution of triphenylphos-
phine (360 mg, 1.374 mmol) in acetonitrile (7 mL) was added
dropwise over 2 min. The mixture was heated under reflux
5,5′-Dib r om o-3,3′-d ica r b oxy-2,2′-d ih yd r oxyd ip h en yl-
m eth a n e (25). 5-Bromosalicylic acid (24, 8.62 g, 40 mmol)
was placed in a 250 mL three-necked, round-bottomed flask
equipped with a mechanical stirrer, a 100 mL pressure-
equalizing dropping funnel, and a thermometer. The solid was

Alkenyldiarylmethanes as Anti-HIV-1 Agents
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 16 3225
dissolved in methanol (10 mL), and the mixture was vigorously
stirred at dry ice-acetone bath temperature while concen-
trated sulfuric acid (70 mL) was added at such a rate to keep
the temperature below 0 °C. The reaction mixture was stirred
on an ice bath for 1 h and then cooled in a dry ice-acetone
bath again. An mixture of 37% aqueous formaldehyde solution
(20 mL) and methanol (20 mL) was added at such a rate to
keep the temperature below 0 °C. The mixture was stirred at
0 °C for 4 h and was left overnight at room temperature. It
was poured on crushed ice (0.5 kg), and the precipitate was
filtered and dried, first at room temperature overnight and
then in a vacuum desiccator, to afford a solid (7.00 g, 78%).
The product was recrystallized from methanol: mp 230 °C dec;
IR (KBr) 3500-2500, 1663, 1607, 1432, 1288, 1238, 1181, 885,
In Vitr o An ti-HIV Assa y. Anti-HIV screening of test
compounds against various viral isolates and cell lines was
performed as previously described.66 This cell-based microtiter
assay quantitates the drug-induced protection from the cyto-
pathic effect of HIV. Data are presented as the percent control
of XTT values for the uninfected, drug-free control. EC50
values reflect the drug concentration that provides 50%
protection from the cytopathic effect of HIV-1 in infected
cultures, while the IC50 reflects the concentration of drug that
causes 50% cell death in the uninfected cultures. XTT-based
results were confirmed by measurement of cell-free superna-
tant reverse transcriptase and p24 levels.
Mech a n istic Assa ys. Binding of HIV-1RF to PBLs was
measured by a p24-based assay.67 Briefly, 2 × 105 CEM-SS
cells were incubated with a concentrated stock of virus for 30
min at 37 °C in the absence or presence of various concentra-
tions of inhibitor, the unbound virus was washed away, and
the cell-associated virus was solubilized in 1% Triton X-100
and 1% BSA and analyzed by the p24 antigen capture assay
as previously described. The effects of inhibitors on the in vitro
activity of purified RT (kind gift of Steve Hughes, NCI-FCRDC,
Frederick, MD) were determined by measurement of incorpo-
ration of [32P]TTP into the poly(rA):oligo(dT) (rAdT) homopoly-
mer or [32P]GTP into the poly(rC):oligo(dG) (rCdG) template/
primer systems. Samples (5 µL) were blotted onto DE81
paper, washed with 5% dibasic sodium phosphate as previously
described,18 and then quantitated on a Packard Matrix 9600
direct beta counter. 3′-Azido-2′,3′-dideoxythymidine 5′-tri-
phosphate (AZTTP) and NSC 629243 (UC38) served as a
positive control for inhibition of RT.
To determine if compounds affected the HIV-1 p7 nucleo-
capsid protein zinc fingers, fluorescence measurements of the
Trp37 residue in the C-terminal zinc finger of the HIV-1 p7
nucleocapsid protein were performed as previously described.68
The p7NC protein was prepared at 20 µg/mL in 10 mM sodium
phosphate buffer (pH 7.0) and treated with 25 µM of test
compound, and then after indicated time intervals the samples
were diluted 1/10 in 10 mM sodium phosphate buffer (pH 7.0)
and the fluorescence intensity was measured. The excitation
and emission wavelengths utilized with the Shimadzu RF5000
spectrofluorimeter were 280 and 351 nm, respectively. The
analytical procedure employed to determine the reagent-
induced inhibition of HIV-1 protease activity has been previ-
ously described.67 Recombinant HIV-1 protease (Bachem
BioScience Inc., King of Prussia, PA) and the substrate (Val-
Ser-Gln-Asn-Tyr-Pro-Ile-Val-Gln-NH2, Multiple Peptide Sys-
tems, San Diego, CA) were utilized to determine the concen-
tration of test compound required to inhibit protease activity
by 50% (IC50). Briefly, HIV-1 protease (14.2 nM final) was
mixed with various concentrations of test compounds in 250
mM potassium phosphate buffer, pH 6.5, 2.5% (v/v) glycerol,
0.5 mM dithiothreitol, 0.5 mM EDTA, and 375 mM ammonium
sulfate, after which the substrate was added (30 nmol) and
the reaction incubated at 37 °C for 30 min. Reactions were
terminated by the addition of 20 µL of a mixture of 8 M
guanidine hydrochloride to 10% trifluoroacetic acid (8:1), and
the reaction products were separated by reverse-phase HPLC
on a Nova-Pak C-18 column. Absorbance was measured at
206 nm, peak areas were quantitated, and the percentage
conversion to product was used to calculate the percentage of
control cleavage in the presence of inhibitors.
1
869, 802 cm-1; H NMR (200 MHz, acetone-d6) δ 7.88 (d, J )
2.4 Hz, 2 H), 7.55 (d, J ) 2.4 Hz, 2 H), 4.00 (s, 2 H); CIMS
m/ e (relative intensity) 447 (MH+, 100), 429 (MH+ - H2O, 84).
Anal. Calcd (C15H10Br2O6) C, H.
5,5′-Dibr om o-2,2′-d im eth oxy-3,3′-bis(m eth oxyca r bon -
yl)d ip h en ylm eth a n e (26). 5,5′-Dibromo-3,3′-dicarboxy-2,2′-
dihydroxydiphenylmethane (25, 7.52 g, 16.86 mmol) was
placed in
equipped with
a
500 mL three-necked, round-bottomed flask
mechanical stirrer, a 50 mL pressure-
a
equalizing dropping funnel, and a reflux condenser connected
to a drying tube. The acid was dissolved in acetone (220 mL,
Mallincrodt, AR), and ground anhydrous potassium carbonate
(19.00 g, 137.5 mmol) was added, followed by dimethyl sulfate
(11.47 g, 8.6 mL, 90.7 mmol). The reaction mixture was stirred
vigorously under reflux for 2 days. The solvent was removed
in vacuo, and water (100 mL) was added, followed by meth-
ylene chloride (80 mL). The resulting mixture was filtered
through a Celite pad, the organic phase was separated, the
residue on the filtration funnel was washed twice with
methylene chloride (2 × 30 mL), and the second filtrate used
for re-extraction of the aqueous phase. The combined organic
extracts were washed with water, dried (sodium sulfate), and
evaporated in vacuo to yield the product as a colorless solid
(8.2 g, 97%): mp 71-72 °C (hexane); IR (KBr) 2947, 1738,
1650, 1577, 1466, 1417, 1320, 1286. 1236, 1194, 997 cm-1; 1H
NMR (500 MHz, CDCl3) δ 7.82 (d, J ) 2.5 Hz, 2 H), 7.32 (d, J
) 2.5, 2 H), 3.99 (s, 2 H), 3.90 (s, 6 H), 3.75 (s, 6 H); CIMS
m/ e (relative intensity) 504 (M+ + 2, 29), 502 (M+, 46), 500
(24), 470 (M+ - MeOH, 100). Anal. (C19H18Br2O6) C, H.
5,5′-Dibr om o-2,2′-d im eth oxy-3,3′-bis(m eth oxyca r bon -
yl)ben zop h en on e (27). 5,5′-Dibromo-2,2′-dimethoxy-3,3′-bis-
(methoxycarbonyl)diphenylmethane (26, 6.34 g, 12.63 mmol)
was placed in a 250 mL one-necked, round-bottomed flask
equipped with a Teflon-coated magnetic stirring bar and a
reflux condenser connected to a drying tube. Acetic anhydride
(128 mL) was added, the solution was cooled in an ice bath,
and chromic anhydride (5.3 g, 5.30 mmol) was added in small
portions over 10 min. The bath was removed, and the mixture
was stirred at room temperature for 2 h, heated under reflux
for 1 min, and then allowed to stand overnight at room
temperature. The chromium salts were filtered off and washed
with methylene chloride (5 × 5 mL). The solvent was removed
in vacuo, and the solidified residue was flash chromatographed
on silica gel (90 g). Elution with methylene chloride afforded
the benzophenone derivative 27 (5.7 g, 88%): mp 128 °C
(methylene chloride-hexane); IR (KBr) 3080, 3000, 2946, 1737,
1648, 1576, 1465, 1417, 1285, 1236, 1194, 1167, 988 cm-1; 1H
NMR (200 MHz, CDCl3) δ 8.08 (d, J ) 2.7 Hz, 2 H), 7.80 (d, J
) 2.7 Hz, 2 H), 3.92 (s, 6 H), 3.57 (s, 6 H); FABMS m/ e (relative
intensity) 517 (MH+, 15), 515 (MH+, 5), 485 (MH+ - MeOH,
28). Anal. (C19H16Br2O7) C, H.
Scr een in g a ga in st HIV-1 Vir u ses Con ta in in g NNRTI
Resista n ce Mu ta tion s. These studies were performed as
previously described.69
5′,5′′-Dibr om o-2′,2′′-d im eth oxy-3′,3′′-bis(m eth oxyca r bo-
n yl)-1,1-d ip h en yl-1-h ep ten e (28). The compound was pre-
pared from 27 as described for the conversion of 6 to 8 to yield
the product an oil in 41% yield: IR (neat) 3071, 2951, 2854,
1723, 1571, 1462, 1437, 1413, 1284, 1195, 1167, 1145, 804, 738,
705 cm-1; 1H NMR (CDCl3, 200 MHz) δ 7.88 (d, J ) 2.6 Hz, 1
H), 7.80 (d, J ) 2.6 Hz, 1 H), 7.56 (d, J ) 2.6 Hz, 1 H), 7.50
(d, J ) 2.6 Hz, 1 H), 6.07 (t, J ) 7.4 Hz, 1 H), 3.88 (s, 3 H),
3.875 (s, 3 H), 3.52 (s, 3 H), 3.48 (s, 3 H), 2.12 (m, 2 H), 1.46
(m, 2 H), 1.28 (m, 4 H), 0.88 (t, J ) 6.9 Hz, 3 H); CIMS m/e
Ack n ow led gm en t. This research was made possible
by NIH Grant RO1-AI-36624 and Contracts NO1-CM-
17513 and NO1-CM-37818.
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