Synthesis and anti-HIV activity of new metabolically stable alkenyldiarylmethane non-nucleoside reverse transcriptase inhibitors incorporating N-methoxy imidoyl halide and 1,2,4-oxadiazole systems

Article abstract of DOI:10.1021/jm070236e

The alkenyldiarylmethanes (ADAMs) are a unique class of non-nucleoside reverse transcriptase inhibitors (NNRTIs) that are capable of inhibiting HIV-1 reverse transcriptase (RT) through an allosteric mechanism. However, the potential usefulness of the ADAM

Full text of DOI:10.1021/jm070236e

3314
J. Med. Chem. 2007, 50, 3314-3321
Synthesis and Anti-HIV Activity of New Metabolically Stable Alkenyldiarylmethane
Non-Nucleoside Reverse Transcriptase Inhibitors Incorporating N-Methoxy Imidoyl Halide and
1,2,4-Oxadiazole Systems
Takeshi Sakamoto,† Matthew D. Cullen,† Tracy L. Hartman,‡ Karen M. Watson,‡ Robert W. Buckheit,‡
Christophe Pannecouque,§ Erik De Clercq,§ and Mark Cushman*,†
Department of Medicinal Chemistry and Molecular Pharmacology, School of Pharmacy and Pharmaceutical Sciences, and
the Purdue Cancer Center, Purdue UniVersity, West Lafayette, Indiana 47907, ImQuest BioSciences, 7340 ExecutiVe Way, Suite R,
Frederick, Maryland 21704, and Rega Institute for Medical Research, Katholieke UniVersiteit LeuVen, B-3000 LeuVen, Belgium
ReceiVed March 1, 2007
The alkenyldiarylmethanes (ADAMs) are a unique class of non-nucleoside reverse transcriptase inhibitors
(NNRTIs) that are capable of inhibiting HIV-1 reverse transcriptase (RT) through an allosteric mechanism.
However, the potential usefulness of the ADAMs is limited by the presence of metabolically labile methyl
ester moieties that are hydrolyzed by nonspecific esterases present in blood plasma, resulting in the formation
of the inactive carboxylic acid metabolites. Therefore, to discover metabolically stable ADAMs, the design
and synthesis of a new class of ADAMs with N-methoxy imidoyl halide and 1,2,4-oxadiazole systems were
attempted. The resulting new ADAM 6 displayed enhanced metabolic stability in rat plasma (t1/2 ) 61 h)
along with the ability to inhibit HIV-1 reverse transcriptase and the cytopathic effect of HIV-1RF and HIV-
1IIIB at submicromolar concentrations.
Introduction
methyl esters and an adjacent methyl ether present in 1.3 ADAM
2 showed very potent anti-HIV-1 activity, with EC50 values of
0.03 µM (HIV-1RF in CEM-SS cells) and 0.09 µM (HIV-1IIIB
in MT-4 cells). Also, ADAM 2 was active against HIV-1 RT,
with an IC50 ) 0.02 µM, establishing it as the most potent RT
inhibitor among ADAM analogues. However, the methyl ester
moieties of the ADAMs are hydrolyzed to biologically inactive
acids by nonspecific esterases present in blood plasma, which
is unfavorable for their potential therapeutic use. Therefore, we
have focused on searching for hydrolytically stable and phar-
macologically active bioisosteres of the methyl esters on the
ADAM phenyl rings and side chain.
The human immunodeficiency virus (HIV), the causative
agent of acquired immunodeficiency syndrome (AIDS), was
identified in the early 1980s. The AIDS epidemic has spread
throughout the world and today an estimated 39.5 million people
are living with HIV/AIDS.1 The potent therapeutic agents that
have been developed to combat the progression of AIDS are
based on four classes of drugs: nucleoside reverse transcriptase
inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibi-
tors (NNRTIs), protease inhibitors (PIs), and fusion inhibitors
(enfuvirtide). Treatment with combinations of these antiviral
agents that target the viral enzymes reverse transcriptase (RT)
and protease, termed highly active antireteroviral therapy
(HAART), has been more successful than monotherapeutic
regimens.2 There are, however, problems with drug toxicity and
multidrug resistance after prolonged therapy. NNRTIs have the
advantage that they are minimally toxic; however, there are
significant problems with the development of NNRTI resistance.
Therefore, the development of new NNRTIs with less severe
side effects and more favorable drug resistance profiles will be
essential to the future management of HIV infection.
Results and Discussion
Design. The N-methoxy imidoyl fluoride and 3-methyl-1,2,4-
oxadiazol-5-yl systems have been previously reported as
metabolically stable bioisosteres for methyl esters in azabicyclic
ring systems.15-17 Examination of electrostatic potential maps
revealed broad similarities between a methyl ester and these
bioisosteres. These results encouraged the design of new ADAM
analogues in which the methyl esters were replaced by N-
methoxy imidoyl fluoride and 3-methyl-1,2,4-oxadiazol-5-yl
bioisosteres.
We envisaged new ADAMs 3 and 6 as metabolically stable
analogues of ADAM 2 with potent anti-HIV-1 activity. In these
ADAMs, the methyl esters on the phenyl rings and side chains
of 1 and 2 are replaced with N-methoxy imidoyl fluoride or
3-methyl-1,2,4-oxadiazol-5-yl systems (Chart 1). In addition,
the new ADAMs 4 and 5 were proposed to examine the effect
of halide size on anti-HIV activity. The synthesis of ADAM 7
was also planned to evaluate the effect of the N-methoxy imidoyl
fluoride replacement on anti-HIV activity by direct comparison
with the ADAM 2.
Among NNRTIs, the alkenyldiarylmethanes (ADAMs) are a
relatively new class of inhibitors that are active at submicromolar
concentrations.3-14 In addition, ADAMs have displayed syn-
ergistic activity with AZT and enhanced activity when tested
against AZT-resistant strains of HIV-1.4,5,7,8 Certain ADAMs,
for example, compounds 1 and 2, have been found to inhibit
HIV-1 RT and the cytopathic effect of HIV-1 in cell culture.
ADAM 1 inhibited HIV-1 RT, with an IC50 value of <1.0 µM
and displayed an EC50 value of 1.0 µM for inhibition of HIV-
1
IIIB in MT-4 cells11,13 (Table 1). The more recently synthesized
ADAM 2 features an oxazolone replacement for one of the
* To whom correspondence should be addressed. Phone: 765-494-1465.
Fax: 765-494-6790. E-mail: cushman@pharmacy.purdue.edu.
† Purdue University.
Chemistry. Our previous work has already established a
general and practical method to synthesize ADAMs using Pd-
catalyzed reactions (Sonogashira reaction, hydrostannation, and
Stille coupling; Scheme 1).12-14 First, the synthesis of ADAM
‡ ImQuest BioSciences.
§ Katholieke Universiteit Leuven.
10.1021/jm070236e CCC: $37.00 © 2007 American Chemical Society
Published on Web 06/19/2007

Anti-HIV ActiVity of Alkenyldiarylmethane
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 14 3315
Table 1. Anti-HIV Activities, Cytotoxicities, and Metabolic Stabilities of ADAM Analoguesa
EC50c (µM)
CC50d (µM)
MT-4 cells
b
IC50
cmpd
(µM)
HIV-1RF
HIV-1IIIB
HIV-2ROD
CEM-SS cells
rat plasma t1/2 ( SD (min)
1
2
1.0
0.02
0.60
0.85
0.67
0.55
0.25
0.02
NAe
NAe
0.7
1.0
0.09
1.2
NAe
NAe
NAe
NAe
NAe
NAe
NAe
NAe
6.0
5.1
1.3
2.1
2.9
3.9
5.2
5.4
6.1
16.86
3.9
13.3
12.4
21.0
16.8
18.4
0.76 ( 0.04
1.30 ( 0.09
4970 ( 795
156 ( 21
3641 ( 14.1
9.2 ( 1.5
NTg
4
5f
6
6.3
0.24
0.29
NAe
NAe
7
22
24
0.5
>100
>100
NAe
NAe
NTg
a All data represent mean values of at least two separate experiments. b Inhibitory activity versus HIV-1 reverse transcriptase with poly(rC).oligo(dG) as
the template primer. c EC50 is the 50% effective concentration for inhibition of cytopathicity of HIV-1RF in CEM-SS cells, HIV-1IIIB in MT-4 cells, or
HIV-2ROD in MT-4 cells. d CC50 is the 50% cytotoxic concentration for the mock-infected CEM-SS cell or MT-4 cells. e Not active. f A mixture (1:1) of E-
and Z-isomers. g Not tested.
Chart 1
Scheme 1
3 was undertaken. The N-methoxy imidoyl fluoride 14 was a
key intermediate corresponding to the general intermediate 8
in Scheme 1. To prepare intermediate 14, two previously
reported methods were applied.17 One was the fluorination of
N-methoxyamide 12 with bis(diethyl)aminosulfur trifluoride
(DAST)17,18 or bis(2-methoxyethyl)aminosulfur trifluoride (Deoxo-
Fluor),19 and the other was the replacement reaction of N-
methoxy imidoyl bromide 13 with calcium fluoride-supported
alkali metal fluorides (KF-CaF2 and CsF-CaF2).17,20 However,
these reactions afforded complex mixtures and attempts to syn-
thesize imidoyl fluoride 14 were all unsuccessful. The difficulty
of the synthesis can probably be attributed to the low nucleo-
philicity and high Lewis basicity of fluoride ions, which are
capable of abstracting the R-hydrogens of imidoyl groups. The

3316 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 14
Sakamoto et al.
Scheme 2a
a Reagents and conditions: (a) PdCl2(PPh3)2, CuI, TEA, THF, room temperature, 12 h; (b) Bu3SnH, Pd(PPh3)4, THF, room temperature, 2 h; (c) LiOH,
dioxane-H2O (6:1), 60 °C, 22 h; (d) H2NOMe, EDCI, DMAP, CH2Cl2, room temperature, 15 h; (e) Pd(PPh3)4, CuI, CsF, DMF, 60 °C, 0.5 h; (f) TFA-
CH2Cl2 (1:1), 0 °C, 3 h; (g) H2NOMe, EDCI, DMAP, CH2Cl2, room temperature, 24 h; (h) PPh3, CCl4, CH3CN, reflux 3.5 h; (i) PPh3, CBr4, CH3CN, reflux,
2 h.
difficulties encountered in the synthesis of ADAM 3 led us to
focus on the imidoyl fluorides 6 and 7, as well as the higher
molecular weight halides 4 and 5.
been previously reported.12,13 The Sonogashira coupling of
starting materials 15 with 16 and the hydrostannation of 17 were
carried out on a large scale. Chemoselective cleavage of methyl
ester 18 with LiOH in dioxane-H2O afforded the corresponding
carboxylic acid 19.22 Subsequent EDCI-promoted coupling of
19 with methoxyamine gave N-methoxyamide 20.
In the next step, the Stille coupling of 20 with 2113 was
attempted using the reaction conditions previously described
by Mee et al.23 The significant feature of their conditions is
that addition of a catalytic amount of cuprous iodide accelerates
the reaction rate in a highly polar solvent such as DMF.
However, in the case of the reaction of 20 with 21,13 no reaction
was observed, probably due to the chelation of the N-
ADAMs 4 and 5 were synthesized as outlined in Scheme 2.
The key functional groups, N-methoxy imidoyl chloride and
bromide, were introduced after the construction of the ADAM
framework to avoid the risk of Pd-catalyzed reactions with the
imidoyl halides. As a related example, Chang et al. have reported
that an N-methoxy imidoyl bromide was successfully employed
as an efficient coupling partner in the Pd-catalyzed Stille
coupling reaction with various organotin compounds.21
The syntheses of intermediates 16 and 21, which are the
precursors of the two aromatic rings of ADAMs 4 and 5, have

Anti-HIV ActiVity of Alkenyldiarylmethane
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 14 3317
Scheme 3a
a Reagents and conditions: (a) DAST, CH2Cl2, 0 °C, 0.5 h; (b) TMOSTf,
TEA, dioxane, room temperature, 2 h; (c) acetamide oxime, TBTU, HOBt,
DIPEA, DMF, room temperature, 0.5 h, then heated at 110 °C, 3 h; (d)
TMSCHN2, toluene-MeOH (2:1), room temperature.
Chart 2
methoxyamide group of 20 and the neighboring methoxy group
to PdII-complexes. In this case, the N-methoxyamide group could
be deprotonated by cesium fluoride and act as a ligand for
palladium.24 To improve this reaction, 1.2 equiv of cuprous
iodide were used. Surprisingly, addition of a large amount of
cuprous iodide afforded the coupling product 22 quantitatively.
The tert-butyl protecting group of 22 was removed with
trifluoroacetic acid to give carboxylic acid 23. EDCI coupling
of 23 with methoxyamine provided di-N-methoxyamide 24.
Finally, chlorination of 24 with PPh3-CCl4 afforded ADAM 4
in 71% yield. Similarly, bromination of 24 with PPh3-CBr4
was attempted. However, the reaction unfortunately afforded a
nearly equimolar mixture of mixture of Z and E isomers of
ADAM 5 in 75% yield. The isomeric mixture was tested for
anti-HIV activity because separation by column chromatography
was unsuccessful. The isomerization of the olefin was probably
caused by Br2 or Br+, which was generated from the decom-
position of [Br3C-PPh3]+Br- complex in refluxing acetonitrile.
The syntheses of ADAMs 6 and 7 were performed according
to Scheme 3. Fluorination of 22 with DAST afforded imidoyl
fluoride 25. Cleavage of the tert-butyl ester of 25 was achieved
using TMSOTf-Et3N to give carboxylic acid 26.25 ADAM 6,
incorporating the 3-methyl-1,2,4-oxadiazole system, was syn-
thesized in 46% yield from carboxylic acid 26 and acetamide
oxime using O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyluro-
nium tetrafluoroborate (TBTU).26 Also, carboxylic acid 26 was
converted into ADAM 7 with (trimethylsilyl)diazomethane in
82% yield.
Figure 1. Electrostatic potentials mapped onto Connolly surfaces of
models 27 (top), 28, 29, 30, and 31 (bottom). Red, electronegative;
blue, electropositive.
electronic character of these methyl ester replacements, com-
pounds 27-31 (Chart 2) were modeled using Sybyl 7.1. The
models were constructed and then energy minimized using the
MMFF94s force field and MMFF94 charges, and the electro-
static potentials were mapped onto the Connolly surfaces. The
results displayed in Figure 1 document a significant variation
in the sizes and electronic characters of these replacements
relative to a methyl ester itself, but in spite of this fact, the
potencies of ADAMs 4, 5, 6, and 7 as inhibitors of HIV-1
reverse transcriptase are all very close, with submicromolar IC50
values. This may reflect the well-known ability of the flexible
NNRTI binding pocket to mold itself to the shape of the
inhibitor.
In the cellular assays for inhibition of the cytopathic effect
of the virus, ADAMs 6 and 7 displayed anti-HIV-1RF activity
in the submicromolar range. Also, the ADAMs 4-7 produced
EC50 values versus HIV-1IIIB between 0.24 and 6.3 µM.
Comparison between ADAMs 2 and 7 indicates that the
N-methoxy imidoyl fluoride system retained the desired anti-
HIV activity, although the potency of 7 was somewhat lower,
both with regard to RT inhibition as well as prevention of the
cytopathic effect of the virus. However, ADAMs 22 and 24,
having N-methoxyamide groups, were inactive both in the
enzymatic and in the cellular tests. Similar to other known
Biological Results and Discussion. Compounds 4-7, 22,
and 24 are a new family of ADAMs characterized by the
presence of N-methoxy imidoyl halide, N-methoxyamide, and
3-methyl-1,2,4-oxadiazole moieties. To visualize the size and

3318 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 14
Sakamoto et al.
(PPh3)2 (0.585 g, 0.817 mmol) was added to the solution. After
stirring for 12 h at room temperature under argon, the mixture was
evaporated and diluted with ethyl acetate (15 mL). The mixture
was filtered through a small pad of silica gel, concentrated, and
purified by column chromatography on silica gel using an ethyl
acetate-hexanes gradient (2-5%) to afford the product 17 (2.23
g, 79%) as a pale yellow oil: IR (Neat) 2973, 1731, 1256, 1215,
1148 cm-1; 1H NMR (300 MHz, CDCl3) δ 7.65 (d, J ) 2.0 Hz, 1
H), 7.34 (d, J ) 2.0 Hz, 1 H), 3.88 (s, 3 H), 3.79 (s, 3 H), 2.34-
2.25 (m, 4 H), 2.25 (s, 3 H), 1.84 (q, J ) 7.2 Hz, 2 H), 1.43 (s, 9
H); 13C NMR (75 MHz, CDCl3) δ 172.43, 166.13, 157.80, 137.77,
132.84, 132.31, 124.41, 119.13, 88.99, 80.24, 79.91, 61.51, 52.16,
34.34, 28.04, 23.99, 18.70, 15.83; ESIMS m/z (rel intensity) 369
(100, MNa+). Anal. Calcd for C20H26O5: C, H.
NNRTIs, all of the ADAMs in this series were inactive against
HIV-2. Regarding metabolic stabilities in rat plasma, the half-
lives of ADAMs 4-6, in which all of the esters were replaced
to N-methoxy imidoyl halide or 3-methyl-1,2,4-oxadiazole
moieties, increased significantly in comparison with the previ-
ously reported ADAMs 1 and 2.
According to previous reports, (Z)-N-methoxy imidoyl halides
isomerize to the E-isomers in HCl-benzene at 40 °C within a
few days.27 However, attempted thermal isomerization of the
Z-isomers in benzene in the absence of HCl at 50 °C failed to
result in any significant isomerization within a period of 1
week.27 Also, in polar protic solvents at elevated temperatures,
imidoyl halides hydrolyze to the corresponding N-methoxy-
amides rather than undergoing isomerization.28 Johnson and
Cornell previously reported that imidoyl halides hydrolyze
thermally at >80 °C in dioxane-water at pH 7.28 In the present
case, neither the NMR spectra of the imidoyl halides nor the
HPLC traces recorded during the hydrolysis studies in rat plasma
provided any evidence for isomerization, indicating that they
are stable under the mild, neutral assay conditions.
The overall results revealed that N-methoxy imidoyl halide,
especially N-methoxy imidoyl fluoride, and 3-methyl-1,2,4-
oxadiazole systems were metabolically stable, biologically active
bioisosteres for methyl esters in the ADAM series. In particular,
ADAM 6 is the best lead compound in this series, having a
2801-fold increase in plasma half-life and maintaining anti-HIV
activities with submicromolar EC50 values. Future studies will
be directed toward modifying the heterocycle on the side chain
of ADAM 6.
(E)-Methyl 5-(6-tert-Butoxy-6-oxo-1-(tributylstannyl)hex-1-
enyl)-2-methoxy-3-methylbenzoate (18). A solution of alkyne 17
(2.55 g, 7.36 mmol) in THF (90 mL) was cooled to 0 °C and
degassed for 10 min with argon, and Pd(PPh3)4 (0.850 g, 0.736
mmol) was added. Tributyltin hydride (2.93 mL, 11.0 mmol) was
added dropwise to the mixture at room temperature and the reaction
mixture was stirred for 2 h under argon. The mixture was evaporated
and diluted with ethyl acetate (50 mL). The mixture was filtered
through a small pad of silica gel, concentrated, and purified by
column chromatography on silica gel using an ethyl acetate-
hexanes gradient (2-4%) to afford the product 18 (4.15 g, 89%)
as a colorless oil: IR (neat) 2928, 2871, 1732, 1148, 1014 cm-1
;
1H NMR (300 MHz, CDCl3) δ 7.16 (d, J ) 2.0 Hz, 1 H), 6.85 (d,
J ) 2.0 Hz, 1 H), 5.70 (t, J ) 6.6 Hz, 1 H), 3.87 (s, 3 H), 3.79 (s,
3 H), 2.26 (s, 3 H), 2.13 (t, J ) 7.3 Hz, 2 H), 1.98-2.08 (m, 2 H),
1.36 (s, 9 H), 1.16-1.68 (m, 14 H), 0.83 (t, J ) 7.4 Hz, 9 H),
0.71-0.96 (m, 6 H); 13C NMR (75 MHz, CDCl3) δ 172.89, 166.83,
155.68, 144.69, 141.52, 140.13, 133.63, 132.09, 127.39, 123.81,
77.86, 61.37, 51.92, 34.98, 29.31, 28.91, 27.94, 27.21, 25.11, 16.04,
13.60, 9.87; ESIMS m/z (rel intensity) 657/659/661 (41/78/100,
MNa+). Anal. Calcd for C32H54O5Sn: C, H.
Experimental Section
Unless noted otherwise, 1H and 13C NMR spectra were recorded
using CDCl3 as the solvent and internal standard at 300 and 75
MHz, respectively. 19F NMR spectra were obtained at 282 MHz in
CDCl3, and chemical shifts are reported in δ values, relative to
CF3CO2H (external standard) at δ 0.00 ppm. IR spectra were
recorded using a Perkin-Elmer 1600 series FT-IR. Flash chroma-
tography was performed with 230-400 mesh silica gel and TLC
was carried out using Baker-flex silica gel IB2-F plates of 0.25
mm thickness. Melting points are uncorrected. Unless otherwise
stated, chemicals and solvents were of reagent grade and used as
obtained from commercial sources without further purification.
Lyophilized rat plasma (lot 065K7555) was obtained from Sigma
Chemical Co., St. Louis, MO. Microanalyses were performed at
the Purdue Microanalysis Laboratory. All yields refer to yields of
isolated compounds.
(E)-5-(6-tert-Butoxy-6-oxo-1-(tributylstannyl)hex-1-enyl)-2-
methoxy-3-methylbenzoic Acid (19). Lithium hydroxide mono-
hydrate (0.328 g, 7.86 mmol) was added to a solution of methyl
ester 18 (4.152 g, 6.513 mmol) in dioxane-H2O (6:1, 60 mL). The
reaction mixture was stirred for 22 h at 60 °C. The mixture was
concentrated in vacuo, acidified with 5% HCl, and diluted with
water (60 mL). The mixture was extracted with ethyl acetate (2 ×
40 mL). The combined organic solvent was washed with brine (2
× 40 mL), dried over sodium sulfate, and concentrated. The residue
was purified by column chromatography on silica gel using 30%
ethyl acetate-hexanes to give the product 19 (2.56 g, 63.0%) as
1
an oil: IR (neat) 2967, 2928, 1732, 1696, 1255, 1149 cm-1; H
NMR (300 MHz, CDCl3) δ 7.78 (br s, 1 H), 7.48 (d, J ) 2.1 Hz,
1 H), 6.94 (d, J ) 2.1 Hz, 1 H), 5.72 (t, J ) 7.4 Hz, 1 H), 3.89 (s,
3 H), 2.31 (s, 3 H), 2.11 (t, J ) 7.4 Hz, 2 H), 2.00 (t, J ) 7.4 Hz,
2 H), 1.61 (quint, J ) 7.4 Hz, 2 H), 1.36 (s, 9 H), 1.14-1.46 (m,
12 H), 0.83 (t, J ) 7.2 Hz, 9 H), 0.71-0.97 (m, 6 H); 13C NMR
(75 MHz, CDCl3) δ 172.89, 167.15, 155.19, 144.30, 141.87, 141.73,
135.23, 131.12, 128.65, 121.48, 79.93, 62.00, 34.95, 29.35, 28.89,
27.94, 26.99, 25.04, 16.47, 13.60, 9.90; ESIMS m/z (rel intensity)
643/645/647 (15/28/35, MNa+); negative ion ESIMS m/z (rel
intensity) 619/621/623 (39/76/100, M - H+). Anal. Calcd for
C31H52O5Sn: C, H.
(E)-tert-Butyl 6-(4-Methoxy-3-(methoxycarbamoyl)-5-meth-
ylphenyl)-6-(tributylstannyl)hex-5-enoate (20). EDCI (2.26 g,
11.8 mmol) was added to a mixture of carboxylic acid 19 (2.45 g,
3.92 mmol), methoxyamine hydrochloride (0.983 g, 11.8 mmol),
4-dimethylaminopyridine (0.240 g, 1.96 mmol), and triethylamine
(2.20 mL, 15.7 mmol) in dichloromethane (50 mL). The reaction
mixture was stirred for 15 h at room temperature. After the reaction
was complete, the mixture was diluted with ethyl acetate (100 mL).
The organic solution was washed with 5% HCl (2 × 30 mL), 5%
aq NaHCO3 (2 × 30 mL), and brine (2 × 30 mL) and dried over
sodium sulfate. After removal of solvent in vacuo, the residue was
purified by column chromatography on silica gel using 30% ethyl
acetate-hexanes to give the product 20 (1.56 g, 61%) as an oil:
tert-Butyl 5-Hexynate (15).29 Di-tert-butyl-dicarbonate (29.2 g,
134 mmol) was added portionwise over 1 h to a solution of
5-hexynoic acid (5.00 g, 44.6 mmol) and 4-dimethylaminopyridine
(0.550 g, 4.46 mmol) in t-BuOH (100 mL), and the reaction mixture
was stirred for 2 h at room temperature. The mixture was evaporated
and diluted with ethyl acetate (80 mL). The organic solvent was
washed with 5% HCl (2 × 30 mL), 5% aq NaHCO3 (2 × 30 mL),
brine (2 × 20 mL), and dried over sodium sulfate. After removal
of solvent in vacuo, the residue was purified by column chroma-
tography on silica gel using an ethyl acetate-hexanes gradient (0-
10%) to afford the product 15 (4.65 g, 62%) as a colorless oil: IR
(neat) 3306, 1730, 1149 cm-1; 1H NMR (300 MHz, CDCl3) δ 2.33
(t, J ) 7.2 Hz, 2 H) 2.25 (dt, J ) 7.2, 2.4 Hz, 2 H), 1.94 (t, J )
2.4 Hz, 1 H), 1.78 (q, J ) 7.2 Hz, 2 H), 1.42 (s, 9 H); 13C NMR
(75 MHz, CDCl3) δ 172.31, 83.38, 80.16, 68.86, 34.11, 28.00,
27.79, 23.72, 21.36, 17.37; GC-CIMS m/z (rel intensity) 169 (3,
MH+), 113 (100). Anal. Calcd for C10H16O2: C, H.
Methyl 5-(5-tert-Butoxycarbonyl-pent-1-ynyl)-2-methoxy-3-
methylbenzonate (17). A solution of alkyne 15 (1.648 g, 9.800
mmol), iodide 16 (2.500 g, 8.167 mmol), cuprous iodide (0.156 g,
0.817 mmol), and triethylamine (2.30 mL, 16.3 mmol) in THF (50
mL) was cooled to 0 °C and degassed for 10 min with argon. PdCl2-

Anti-HIV ActiVity of Alkenyldiarylmethane
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 14 3319
IR (neat) 3340, 1730, 1683, 1464, 1148 cm-1; 1H NMR (300 MHz,
CDCl3) δ 10.15 (br s, 1 H), 7.37 (d, J ) 2.3 Hz, 1 H), 6.82 (d, J
) 2.3 Hz, 1 H), 5.70 (t, J ) 7.4 Hz, 1 H), 3.87 (s, 3 H), 3.75 (s,
3 H), 2.26 (s, 3 H), 2.12 (t, J ) 7.4 Hz, 2 H), 2.10 (q, J ) 7.4 Hz,
2 H), 1.60 (quint, J ) 7.4 Hz, 2 H), 1.36 (s, 9 H), 1.14-1.46 (m,
12 H), 0.83 (t, J ) 7.2 Hz, 9 H), 0.71-0.95 (m, 6 H); 13C NMR
(75 MHz, CDCl3) δ 172.99, 164.37, 153.38, 144.62, 141.56, 133.37,
130.85, 127.23, 124.03, 79.93, 64.31, 61.37, 34.97, 29.36, 28.90,
27.95, 27.20, 25.08, 15.95, 13.62, 9.89; ESIMS m/z (rel intensity)
672/674/676 (42/94/100, MNa+). Anal. Calcd for C32H55NO5Sn:
C, H, N.
(E)-tert-Butyl 6-(3,7-Dimethyl-2-oxo-2,3-dihydrobenzo[d]ox-
azol-5-yl)-6-(4-methoxy-3-(methoxycarbamoyl)-5-methylphenyl)-
hex-5-enoate (22). A mixture of aryl iodide 2113 (56 mg, 0.188
mmol), organostannane 20 (147 mg, 0.225 mmol), and cesium
fluoride (104 mg, 0.675 mmol) in DMF (5 mL) was cooled to 0
°C and degassed for 10 min with argon. Pd(PPh3)4 (22 mg, 0.019
mmol) and cuprous iodide (43 mg, 0.255 mmol) were added to the
mixture. The reaction mixture was stirred for 0.5 h at 60 °C under
argon. After the reaction was complete, the reaction mixture was
diluted with dichloromethane (20 mL) and water (10 mL). After
vigorous shaking, the mixture was filtered through celite with ethyl
acetate (80 mL). The organic layer was separated, washed with
brine (3 × 30 mL), dried over sodium sulfate, and concentrated.
The residue was purified by column chromatography on silica gel
using 30% ethyl acetate-hexanes to give the product 22 (100 mg,
100%) as an oil: IR (neat) 3308, 2975, 2934, 1777, 1726, 1674,
1470, 1151, 1001, 750 cm-1; 1H NMR (300 MHz, CDCl3) δ 10.02
(br s, 1 H), 7.61 (d, J ) 1.2 Hz, 1 H), 7.05 (d, J ) 1.2 Hz, 1 H),
6.70 (d, J ) 1.0 Hz, 1 H), 6.55 (d, J ) 1.0 Hz, 1 H), 5.92 (t, J )
7.5 Hz, 1 H), 3.88 (s, 3 H), 3.83 (s, 3 H), 3.32 (s, 3 H), 2.29 (s, 6
H), 2.20 (t, J ) 7.5 Hz, 2 H), 2.08 (q, J ) 7.5 Hz, 2 H), 1.70
(quint, J ) 7.5 Hz, 2 H), 1.37 (s, 9 H); 13C NMR (75 MHz, CDCl3)
δ 172.81, 164.10, 155.15, 154.97, 140.59, 140.40, 138.76, 136.33,
135.98, 131.54, 131.16, 129.98, 124.52, 123.62, 119.77, 104.60,
80.11, 64.33, 61.37, 34.95, 29.14, 28.13, 27.93, 25.13, 16.00, 14.36;
ESIMS m/z (rel intensity) 547 (100, MNa+); negative ion ESIMS
m/z (rel intensity) 523 [100, (M - H+)-]. Anal. Calcd for C29H36-
NO7: C, H, N.
washed with 2% HCl (2 × 15 mL), 5% aq NaHCO3 (2 × 15 mL),
and brine (2 × 15 mL) and dried over sodium sulfate. After removal
of solvent in vacuo, the residue was purified by column chroma-
tography on silica gel using 3% methanol-ethyl acetate to give
the product 24 (40 mg, 53%) as an oil: IR (neat) 3435, 1772, 1648,
1
1471, 1064 cm-1; H NMR (300 MHz, CDCl3) δ 10.34 (br s, 1
H), 8.76 (br s, 1 H), 7.61 (s, 1 H), 7.02 (s, 1 H), 6.69 (s, 1 H), 6.56
(s, 1 H), 5.94 (t, J ) 7.6 Hz, 1 H), 3.89 (s, 3 H), 3.83 (s, 3 H), 3.67
(s, 3 H), 3.31 (s, 3 H), 2.28 (s, 6 H), 2.14-1.99 (m, 4 H), 1.73-
1.87 (m, 2 H); 13C NMR (75 MHz, CDCl3) δ 170.40, 164.30,
155.25, 154.97, 140.65, 140.36, 138.60, 136.06, 135.93, 131.80,
131.12, 129.80, 129.59, 124.43, 123.56, 119.73, 104.58, 64.21,
63.90, 61.32, 32.31, 29.11, 28.10, 25.26, 15.97, 14.30; ESIMS m/z
(rel intensity) 498 (100, MH+), 520 (70, MNa+); negative ion
ESIMS m/z (rel intensity) 496 [86, (M -H+)-]. Anal. Calcd for
C26H31N3O7: C, H, N.
(Z)-5-((1E,6Z)-6-Chloro-1-(3,7-dimethyl-2-oxo-2,3-dihydroben-
zo[d]oxazol-5-yl)-6-(methoxyimino)hex-1-enyl)-N,2-dimethoxy-
3-methylbenzimidoyl Chloride (4). Carbon tetrachloride (0.16 mL,
1.61 mmol) was added to a solution of di-N-methoxyamide 24 (40
mg, 0.080 mmol) and triphenylphosphine (84 mg, 0.322 mmol) in
acetonitrile (5 mL). The reaction mixture was stirred for 0.5 h at
room temperature and for 3.5 h at reflux. Additional triphenylphos-
phine (42 mg, 0.161 mol) and carbon tetrachloride (0.1 mL, 1.01
mmol) were added, and the mixture was stirred for 1 h at reflux.
After the reaction was complete, the solvent was evaporated in
vacuo. The residue was purified by column chromatography on
silica gel using 20% ethyl acetate-hexanes to give the product 4
(30 mg, 71%) as an oil: IR (neat) 2938, 1779, 1464, 1025 cm-1
;
1H NMR (300 MHz, CDCl3) δ 7.08 (d, J ) 2.0 Hz, 1 H), 6.99 (d,
J ) 2.0 Hz, 1 H), 6.76 (d, J ) 1.1 Hz, 1 H), 6.57 (d, J ) 1.1 Hz,
1 H), 5.93 (t, J ) 7.4 Hz, 1 H), 4.07 (s, 3 H), 3.88 (s, 3 H), 3.83
(s, 3 H), 3.33 (s, 3 H), 2.47 (t, J ) 7.4 Hz, 2 H), 2.31 (s, 3 H), 2.29
(s, 3 H), 2.16 (q, J ) 7.4 Hz, 2 H), 1.79 (quint, J ) 7.4 Hz, 2 H);
ESIMS m/z (rel intensity) 556/558/560 (100/65/11, MNa+). Anal.
Calcd for C26H29Cl2N3O5: C, H, N.
Stereoisomeric Mixture of (Z)-5-((1E,6Z)-6-Bromo-1-(3,7-
dimethyl-2-oxo-2,3-dihydrobenzo[d]oxazol-5-yl)-6-(methoxyimi-
no)hex-1-enyl)-N,2-demethoxy-3-methylbenzimidoyl Bromide
(E-5) and (Z)-5-((1Z,6Z)-6-Bromo-1-(3,7-dimethyl-2-oxo-2,3-
dihydrobenzo[d]oxazol-5-yl)-6-(methoxyimino)hex-1-enyl)-N,2-
demethoxy-3-methylbenzimidoyl Bromide (Z-5). Carbon tetra-
bromide (893 mg, 2.47 mmol) was added to a solution of di-N-
methoxyamide 24 (123 mg, 0.247 mmol) and triphenylphosphine
(342 mg, 1.24 mmol) in acetonitrile (15 mL). The reaction mixture
was stirred for 0.5 h at room temperature and for 2 h at reflux. The
solvent was evaporated in vacuo and the residue was purified by
column chromatography on silica gel using 25% ethyl acetate-
hexanes to give an equimolar mixture of E and Z isomers of ADAM
5 (115 mg, 75%) as an oil: IR (neat) 2936, 1778, 1757, 1640,
(E)-6-(3,7-Dimethyl-2-oxo-2,3-dihydrobenzo[d]oxazol-5-yl)-6-
(4-methoxy-3-(methoxycarbamoyl)-5-methylphenyl)hex-5-eno-
ic Acid (23). Trifluoroacetic acid (2 mL) was added to a solution
of the tert-butyl ester 22 (99 mg, 0.189 mmol) in dichloromethane
(2 mL). The reaction mixture was stirred for 3 h at 0 °C. After the
reaction was complete, the solvent was evaporated at room
temperature and the residue was diluted with ethyl acetate (40 mL).
The organic solution was washed with brine (2 × 20 mL) and dried
over sodium sulfate. After removal of solvent in vacuo, the residue
was purified by column chromatography on silica gel using 75%
ethyl acetate-hexanes to give the product 23 (71 mg, 80%) as an
1
1
oil: IR (neat) 3271, 1771, 1645, 1471, 731 cm-1; H NMR (300
1469, 1063 cm-1; H NMR (300 MHz, CDCl3) δ 7.05 (s, 2 H),
MHz, CDCl3) δ 10.22 (s, 1 H), 8.65 (br s, 1 H), 7.63 (d, J ) 1.6
Hz, 1 H), 7.04 (d, J ) 1.6 Hz, 1 H), 6.69 (d, J ) 1.0 Hz, 1 H),
6.56 (d, J ) 1.0 Hz, 1 H), 5.92 (t, J ) 7.4 Hz, 1 H), 3.86 (s, 3 H),
3.82 (s, 3 H), 3.31 (s, 3 H), 2.32 (t, J ) 7.4 Hz, 2 H), 2.28 (s, 6 H),
2.12 (q, J ) 7.4 Hz, 2 H), 1.76 (quint, J ) 7.4 Hz, 2 H); 13C NMR
(75 MHz, CDCl3) δ 177.82, 164.12, 155.09, 154.95, 140.74, 140.33,
138.62, 136.24, 136.05, 131.57, 131.07, 129.95, 129.61, 124.19,
123.55, 119.70, 104.55, 64.24, 61.31, 33.18, 28.84, 28.05, 24.55,
15.90, 14.26; ESIMS m/z (rel intensity) 491 (100, MNa+); negative
ion ESIMS m/z (rel intensity) 467 [100, (M - H+)-]. Anal. Calcd
for C25H28N2O7: C, H, N.
(E)-5-(1-(3,7-Dimethyl-2-oxo-2,3-dihydrobenzo[d]oxazol-5-yl)-
6-(methoxyamino)-6-oxohex-1-enyl)-N,2-dimethoxy-3-methyl-
benzamide (24). EDCI (58 mg, 0.304 mmol) was added to a
solution of carboxylic acid 23 (71 mg, 0.152 mmol), methoxyamine
hydrochloride (25 mg, 0.304 mmol), 4-dimethylaminopyridine (4
mg, 0.030 mmol), and triethylamine (0.06 mL, 0.456 mmol) in
dichloromethane (4 mL). The reaction mixture was stirred for 24
h at room temperature. After the reaction was complete, the mixture
was diluted with ethyl acetate (40 mL). The organic solution was
7.00 (s, 1 H), 6.97 (s, 1 H), 6.77 (s, 1 H), 6.71 (s, 1 H), 6.58 (s, 1
H), 6.52 (s, 1 H), 5.99 (t, J ) 7.4 Hz, 1 H), 5.93 (t, J ) 7.5 Hz, 1
H), 4.11 (s, 3 H), 4.09 (s, 3 H), 3.91 (s, 3 H), 3.88 (s, 3 H), 3.82
(s, 3 H), 3.76 (s, 3 H), 3.35 (s, 3 H), 3.33 (s, 3 H), 2.94-2.64 (m,
4 H), 2.38 (s, 3 H), 2.31 (s, 3 H), 2.29 (s, 3 H), 2.23 (s, 3 H),
2.22-2.05 (m, 4 H), 1.87-1.69 (m, 4 H); ESIMS m/z (rel intensity)
644/646/648 (47/100/49, MNa+).
(E)-tert-Butyl 6-(3,7-Dimethyl-2-oxo-2,3-dihydrobenzo[d]ox-
azol-5-yl)-6-(3-((Z)-fluoro(methoxyimino)methyl)-4-methoxy-5-
methylphenyl)hex-5-enoate (25). (Diethylamino)sulfur trifluoride
(0.07 mL, 0.510 mmol) was added to a solution of N-methoxyamide
22 (134 mg, 0.255 mmol) in dichloromethane (7 mL) at 0 °C under
argon, and the mixture was stirred for 0.5 h. After quenching the
reaction with 5% aq NaHCO3, the mixture was extracted with ethyl
acetate (2 × 25 mL). The combined organic solvent was washed
with brine (2 × 25 mL) and dried over sodium sulfate. After
removal of solvent in vacuo, the residue was purified by column
chromatography on silica gel using an ethyl acetate-hexanes
gradient (10-13%) to give the product 25 (80 mg, 60%) as an oil:
IR (neat) 2939, 1779, 1727, 1472, 1149, 1051 cm-1; 1H NMR (300

3320 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 14
Sakamoto et al.
1
MHz, CDCl3) δ 7.19 (d, J ) 2.0 Hz, 1 H), 7.04 (s, 1 H), 6.73 (s,
1 H), 6.56 (s, 1 H), 5.93 (t, J ) 7.4 Hz, 1 H), 3.94 (s, 3 H), 3.84
(s, 3 H), 3.32 (s, 3 H), 2.31 (s, 3 H), 2.29 (s, 3 H), 2.21 (t, J ) 7.5
Hz, 2 H), 2.12 (t, J ) 7.5 Hz, 2 H), 1.72 (quint, J ) 7.5 Hz, 2 H),
1.38 (s, 9 H); 13C NMR (75 MHz, CDCl3) δ 170.70, 156.16 (d, J
) 75.2 Hz), 151.65, 147.35, 140.46, 138.72, 135.40, 132.56, 131.22,
129.97, 128.76, 128.75, 123.58, 120.22, 119.84, 104.56, 80.07,
63.06, 60.92, 35.00, 29.16, 28.15, 27.96, 25.19, 16.09, 14.40; 19F
NMR (282 MHz, CDCl3) δ 10.95 (s, 1 F); ESIMS m/z (rel intensity)
494 (100, M-methanol), 495 (30). Anal. Calcd for C29H35FN2O5:
C, H, N.
an oil: IR (neat) 2944, 1778, 1737, 1471, 1050 cm-1; H NMR
(300 MHz, CDCl3) δ 7.18 (d, J ) 1.8 Hz, 1 H), 7.03 (d, J ) 1.8
Hz, 1 H), 6.72 (s, 1 H), 6.55 (s, 1 H), 5.92 (t, J ) 7.4 Hz, 1 H),
3.94 (s, 3 H), 3.84 (s, 3 H), 3.61 (s, 3 H), 3.32 (s, 3 H), 2.32 (s, 3
H), 2.29 (s, 3 H), 2.28 (t, J ) 7.4 Hz, 2 H), 2.12 (q, J ) 7.4 Hz,
2 H), 1.77 (quint, J ) 7.4 Hz, 2 H); 13C NMR (75 MHz, CDCl3)
δ 173.74, 155.56 (d, J ) 93.2 Hz), 151.62, 147.31, 140.70, 140.46,
138.64, 135.35, 132.57, 131.22, 129.64, 128.70, 123.55, 120.20,
119.89, 104.53, 63.04, 60.92, 51.41, 33.38, 29.90, 28.13, 24.92,
16.08, 14.38; 19F NMR (282 MHz, CDCl3) δ 10.86 (s, 1 F); ESIMS
m/z (rel intensity) 485 (100, MH+). Anal. Calcd for C26H29FN2O6:
C, H, N.
(E)-6-(3,7-Dimethyl-2-oxo-2,3-dihydrobenzo[d]oxazol-5-yl)-6-
(3-((Z)-fluoro(methoxy-imino)methyl)-4-methoxy-5-methylphen-
yl)hex-5-enoic Acid (26). Trimethylsilyl trifluoromethanesulfonate
(0.59 mL, 3.27 mmol) was added to a solution of tert-butyl ester
25 (862 mg, 1.64 mmol) and triethylamine (0.46 mL, 3.27 mmol)
in dioxane (25 mL) at room temperature under argon. The reaction
mixture was stirred for 2 h at room temperature. After the reaction
was complete, water (50 mL) was added to the mixture. The mixture
was extracted with ethyl acetate (2 × 35 mL). The combined
organic solvent was washed with brine (2 × 35 mL) and dried
over sodium sulfate. After removal of solvent in vacuo, the residue
was purified by column chromatography on silica gel using an ethyl
acetate-hexanes gradient (30-70%) to give the product 26 (626
mg, 81%) as an oil: IR (neat) 3200, 2941, 1778, 1708, 1471, 1051
RT Inhibition Assay. Inhibition of purified recombinant reverse
transcriptase was measured by the incorporation of [32P]GTP into
poly(rC)/oligo(dG) (rCdG) homopolymer template primers, as
previously described.4,30
In Vitro Antiviral Assays. Evaluation of the antiviral activity
of compounds against HIV-1RF infection in CEM-SS cells was
performed using the XTT cytoprotection assay, as previously
described.30 Evaluation of the antiviral activity of the compounds
against HIV-1 strain IIIB and HIV-2 strain (ROD) in MT-4 cells
was performed using the MTT assay, as previously described.31,32
In Vitro Hydrolytic Stability Study in Rat Plasma. The
alkenyldiarylmethanes 4-7 (with 1,1-diphenylethylene or ben-
zophenone as internal standards) were tested for their hydrolytic
stability, utilizing rat plasma and in vitro methods, as previously
described.11
1
cm-1; H NMR (300 MHz, CDCl3) δ 7.19 (d, J ) 1.4 Hz, 1 H),
7.03 (d, J ) 1.4 Hz, 1 H), 6.73 (d, J ) 1.1 Hz, 1 H), 6.55 (d, J )
1.1 Hz, 1 H), 5.93 (t, J ) 7.4 Hz, 1 H), 3.94 (s, 3 H), 3.84 (s, 3 H),
3.32 (s, 3 H), 2.34 (t, J ) 7.3 Hz, 2 H), 2.30 (s, 3 H), 2.29 (s, 3 H),
2.15 (q, J ) 7.3 Hz, 2 H), 1.77 (quint, J ) 7.3 Hz, 2 H); 13C NMR
(75 MHz, CDCl3) δ 179.02, 155.67 (J ) 89.9 Hz,), 151.80, 147.80,
140.94, 140.40, 138.64, 135.35, 135.33, 132.69, 131.27, 129.47,
128.71, 123.62, 120.10, 119.97, 140.85, 63.12, 60.98, 33.35, 29.06,
28.21, 24.69, 16.14, 14.46; 19F NMR (282 MHz, CDCl3) δ 11.29;
ESIMS m/z (rel intensity) 470.97 (100, MH+); negative ion ESIMS
m/z (rel intensity) 469 [100, (M - H+)-]. Anal. Calcd for C25H27-
FN2O5: C, H, N.
Acknowledgment. This investigation was made possible by
Grant RO1-AI-43637, awarded by the National Institutes of
Health, DHHS. This research was conducted in a facility
constructed with support from Research Facilities Improvement
Program Grant Number C06-14499 from the National Center
for Research Resources of the National Institutes of Health.
Supporting Information Available: HPLC analysis results of
compound 5 and elemental analysis data for all other compounds.
This material is available free of charge via the Internet at http://
pubs.acs.org.
(Z)-5-((E)-1-(3,7-Dimethyl-2-oxo-2,3-dihydrobenzo[d]oxazol-
5-yl)-5-(3-methyl-1,2,4-oxadiazol-5-yl)pent-1-enyl)-N,2-dimethoxy-
3-methylbenzimidoyl Fluoride (6). TBTU (O-(benzotriazol-1-yl)-
N,N,N′,N′-tetramethyluronium tetrafluoroborate; 71 mg, 0.213
mmol) was added to a mixture of carboxylic acid 26 (100 mg, 0.213
mmol), acetamide oxime (16 mg, 0.213 mmol), HOBt (6 mg, 0.043
mmol), and DIPEA (0.19 mL, 1.07 mmol) in DMF (4 mL). The
mixture was stirred for 0.5 h at room temperature and for 3 h at
110 °C. The mixture was diluted with ethyl acetate (50 mL), and
the organic solution was washed with brine (4 × 20 mL), dried
over sodium sulfate, and concentrated. The residue was purified
by preparative TLC using 50% ethyl acetate-hexanes to give the
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product 6 (50 mg, 46%) as an oil. IR (neat) 2956, 1778, 1051 cm-1
;
1H NMR (300 MHz, CDCl3) δ 7.18 (d, J ) 2.0 Hz, 1 H), 7.10 (d,
J ) 2.0 Hz, 1 H), 6.71 (d, J ) 0.8 Hz, 1 H), 6.54 (d, J ) 0.8 Hz,
1 H), 5.93 (t, J ) 7.4 Hz, 1 H), 3.94 (s, 3 H), 3.85 (s, 3 H), 3.32
(s, 3 H), 2.83 (t, J ) 7.7 Hz, 2 H), 2.33 (s, 3 H), 2.31 (s, 3 H), 2.29
(s, 3 H), 2.21 (q, J ) 7.4 Hz, 2 H), 1.95 (quint, J ) 7.5 Hz, 2 H);
13C NMR (75 MHz, CDCl3) δ 179.12, 166.91, 155.61 (d, J ) 98.3
Hz), 151.60, 147.30, 141.34, 140.56, 138.44, 135.25, 135.16,
132.72, 131.26, 128.76, 128.65, 123.56, 120.28, 119.92, 104.52,
63.10, 60.95, 28.93, 28.17, 26.47, 16.13, 14.42, 11.44; 19F NMR
(282 MHz, CDCl3) δ 10.99 (s, 1 F); ESIMS m/z (rel intensity) 509
(100, MH+). Anal. Calcd for C27H29FN4O5: C, H, N.
(E)-Methyl 6-(3,7-Dimethyl-2-oxo-2,3-dihydrobenzo[d]oxazol-
5-yl)-6-(3-(Z)-fluoro-(methoxyimino)methyl)-4-methoxy-5-meth-
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of 2 M solution in hexanes, 0.340 mmol) was added to a solution
of the carboxylic acid 26 (80 mg, 0.170 mmol) in toluene-methanol
(2:1, 4.5 mL). After stirring for 10 min at room temperature, the
excess (trimethylsilyl)diazomethane was quenched by dropwise
addition of acetic acid. The solvent was evaporated, and the residue
was purified by column chromatography using an ethyl acetate-
hexanes gradient (20-40%) to give the product 7 (67 mg, 82%) as

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