Syntheses and bioactivities of tricyclic pyrones

Article abstract of DOI:10.1016/S0040-4020(03)00687-2

In search of compounds that ameliorate the toxicity of amyloid-β (Aβ) peptides, new derivatives of tricyclic pyrones (1-7) were synthesized and their biological activities evaluated. The carboxylic ester and amide derivatives 1-4 were synthesized from a selective carboxylation of C3 methyl of (5aS,7S)-{7-Isopropenyl-3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-oxopyrano [4,3-b][1]benzopyran (8) with LDA followed by benzyl chloroformate or carbon dioxide to provide ester 1 and carboxylic acid 9, respectively. Three isomeric tricyclic pyrone, 5-7, containing adenine moiety at C7 side chain were synthesized from the alkylation of mesylate 13 with adenine, and displacement of chloropurine 15 with amine 14. Although C3-benzyloxycarbonylmethyl analogs 1-3 have marginal ACAT and CETP activities, their modified aspartate analog 4 and C3-methyl-C7-(N3-adeninyl)-2-propyl analog 6 show a significant effect in protecting against neuron-cell death from the toxicity of intracellular accumulation of Aβ or Aβ-containing C-terminal fragments (CTF) of amyloid β precursor protein (APP). N9-Adenine analog 5 is 20-fold less effective than N3-adenine derivative 6 in the protection of neuron-cell death induced by Aβ, while N10-adenine analog 7 was inactive. As a result of this study, compounds 4 and 6 will well serve as lead compounds for further studies of the mechanism of action of Aβ-and CTF-induced neuron-cell death, studies which should enhance the future development of new drugs for the prevention and treatment of AD.

Full text of DOI:10.1016/S0040-4020(03)00687-2

Tetrahedron 59 (2003) 4795–4803
Syntheses and bioactivities of tricyclic pyrones
a
a
a
a
Duy H. Hua,a Xiaodong Huang, Masafumi Tamura, Yi Chen, Melissa Woltkamp,
Lee-Way Jin,b Elisabeth M. Perchellet,c Jean-Pierre Perchellet,c Peter K. Chiang,d
Ichiji Namatamee and Hiroshi Tomodae
,
*
aDepartment of Chemistry, Willard Hall, Kansas State University, Manhattan, KS 66506 USA
bDepartment of Pathology, University of Washington, Seattle, WA 98195-7470 USA
cAnti-Cancer Drug Laboratory, Division of Biology, Ackert Hall, Kansas State University, Manhattan, KS 66506 USA
dDepartment of Applied Biochemistry, Walter Reed Army Institute of Research, Washington, DC 20307-5100 USA
eKitasato Institute for Life Sciences, Kitasato University, 5-9-1, Shirokane, Minato-Ku, Tokyo 108-8641 Japan
Received 24 February 2003; revised 30 April 2003; accepted 30 April 2003
Abstract—In search of compounds that ameliorate the toxicity of amyloid-b (Ab) peptides, new derivatives of tricyclic pyrones (1–7) were
synthesized and their biological activities evaluated. The carboxylic ester and amide derivatives 1–4 were synthesized from a selective
carboxylation of C3 methyl of (5aS,7S)-{7-Isopropenyl-3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-oxopyrano[4,3-b][1]benzopyran (8) with
LDA followed by benzyl chloroformate or carbon dioxide to provide ester 1 and carboxylic acid 9, respectively. Three isomeric tricyclic
pyrone, 5–7, containing adenine moiety at C7 side chain were synthesized from the alkylation of mesylate 13 with adenine, and displacement
of chloropurine 15 with amine 14. Although C3-benzyloxycarbonylmethyl analogs 1–3 have marginal ACAT and CETP activities, their
modified aspartate analog 4 and C3-methyl-C7-(N3-adeninyl)-2-propyl analog 6 show a significant effect in protecting against neuron-cell
death from the toxicity of intracellular accumulation of Ab or Ab-containing C-terminal fragments (CTF) of amyloid b precursor protein
(APP). N9-Adenine analog 5 is 20-fold less effective than N3-adenine derivative 6 in the protection of neuron-cell death induced by Ab,
while N10-adenine analog 7 was inactive. As a result of this study, compounds 4 and 6 will well serve as lead compounds for further studies
of the mechanism of action of Ab-and CTF-induced neuron-cell death, studies which should enhance the future development of new drugs
for the prevention and treatment of AD. q 2003 Elsevier Science Ltd. All rights reserved.
1. Introduction
biological activities of new TPs (Fig. 1) including ACAT
and CETP (cholesteryl ester transfer protein) inhibitory
activities, anticancer activity, and the rescue of cultured
APP C99-expressing neurons from cell death.
The pathogenetic events leading to Alzheimer’s disease
(AD) may reside in the production and deposition of
amyloid-b (Ab) peptides.1 Ab peptides are 39–42-amino
acid hydrophobic polypeptides derived from a trans-
membrane glycoprotein, amyloid b precursor protein
(APP). Based on the bioactivities of pyripyropene A,2 a
potent ACAT (acyl-coenzyme A: cholesterol O-acyltrans-
ferase inhibitor, and arisugacin,3 an acetylcholinesterase
inhibitor, a number of tricyclic pyrone analogs was
synthesized and their bio-activities were studied.4,5 A recent
report on the modulating effect of ACAT on the generation
of Ab peptides6 supports such a possibility, that tricyclic
pyrone (TP) analogs might prevent Ab-induced cellular
toxicity. Those encouraging results5 prompted us to prepare
a number of new derivatives of tricyclic pyrones containing
carboxyl, aspartate, and adenine moieties. We have also
investigated the unusual reactivity of adenine towards alkyl
methanesulfonate. Furthermore, we have determined the
2. Results and discussion
2.1. Syntheses
Based on the synthesis of an ACAT inhibitor, pyripyropene
A (a tetracyclic pyranopyrone),2 several tricyclic pyrano-
pyrones containing various functionalities (Fig. 1) were
synthesized. A common intermediate, compound 8 (an
optically pure compound),4 was used in the synthesis of
1,7 (Schemes 1 and 2) by selective functionalizations of
C3 and C7 side chains. A regioselective deprotonation of the
C3 methyl group of TP 8 can be carried out with a strong
base such as lithium diisopropylamide (LDA) or n-BuLi.
Hence, treatment of 8 with LDA followed by benzyl
chloroformate gave a 46% yield (based on reacted 8) of 1
with 37% recovery of 8 (Scheme 1). Apparently, the anion
of 8 deprotonates a-CH of the benzyl ester function of
product 1 to give 8 and anion of 1. However, in the reaction
Keywords: Alzheimer’s disease; amyloid-b peptides; ACAT inhibitors;
CETP inhibitors; tricyclic pyrones; APP C99-induced cell death.
*
Corresponding author. Tel.: þ1-785-532-6699; fax: þ1-785-532-6666;
e-mail: duy@ksu.edu
0040–4020/03/$ - see front matter q 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0040-4020(03)00687-2

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D. H. Hua et al. / Tetrahedron 59 (2003) 4795–4803
Figure 1.
with carbon dioxide (vide infra), this deprotonation by the
anion of 8 is precluded by its faster reaction with the carbon
dioxide. Chemo-selective hydroboration of 1 with 1 equiv.
of BH3·THF at 2258C for 14 h followed by oxidation with
0.5% NaOH/30% H2O2 at 08C gave a 69% yield (based on
reacted 1) of alcohols 2 along with 26% recovery of 1. The
less hindered C13 double bond was selectively reacted in
the presence of C9a double bond. Based on the 13C NMR
spectrum of 2, two diastereomers at C13 were formed in a
ratio of 1:1 and were inseparable by silica gel column
chromatography and HPLC (normal phase; 2, 3, 5, and 6).
Hence, mixtures of these diastereomers were used for
biological evaluation. A dihydroxylation was resulted when
the hydroboration reaction was carried out at 08C. Yields of
32% of 3 and 14% of 2 were isolated. Again, column
chromatography afforded only an inseparable mixture of
two C13 diastereomers, 3. No other stereoisomers at C9a
and C10 of 3 were detected. The C9a,10 stereochemistry of
3 was tentatively assigned assuming that borane approaches
the C9a–C10 double bond of 2 from the less hindered a face
(same side as C5a–H). Amide 4 was synthesized in 90%
overall yield from pyrone 8 by (1) treatment with LDA in
THF followed by carboxylation with carbon dioxide to give
acid 9 (92% yield), and (2) coupling of 9 with amine 10 in a
solution of N,N0-dicyclohexylcarbodiimide (DCC) in
dichloromethane followed by basic hydrolysis with aqueous
NaOH. Amine 10 was prepared in 85% overall yield from
the coupling of N-(t-butyloxycarbonyl)-L-aspartic acid
4-benzyl ester (11) with p-nitroaniline in a solution of
DCC in dichloromethane followed by acidic hydrolysis of
the t-butoxycarbonyl protecting group with HCl.
Since the isopropenyl group of TPs can be selectively
hydroxylated (vide supra), functional group transformation
of the resulting hydroxyl group would provide a number of
TP analogs. Hence, treatment of 8 with 1 equiv. of BH3·THF
at 2258C followed by oxidation with NaOH and H2O2 gave
an 82% yield (based on reacted 8) of an inseparable mixture
of two C-12 diastereomers, 12, and a 26% recovery of 8
(Scheme 2). 13C NMR spectrum of 12 revealed the two
diastereomers to be present in equal amounts. Mesylation of
12 with triethylamine and methanesulfonyl chloride gave
mesylate 13 in a 94% yield. Treatment of mesylate 13 with
adenine and sodium hydride in DMF at 808C for 20 h
produced 5 and 6 in a ratio of 4:1. Compounds 5 and 6 were
separated by sili0ca gel column chromatography. The
corresponding N7 -analog was not detected under these
reaction conditions. The regiochemistry of these two
isomeric products was assigned0 based on the comparison
of the 1H NMR resonances of C2 H and C80H of the adenine
moiety with the corresponding values reported.7 Moreover,
2D NOESY experiments showed a correlation between

D. H. Hua et al. / Tetrahedron 59 (2003) 4795–4803
4797
Scheme 1.
C80-H (d 7.78 ppm) and CH2N (d 4.01 ppm) of 5 and
between C80-H (d 7.98 ppm) and CH2N (d 4.08 ppm) of 6.
toluene under reflux (25% yield) followed by removal of the
protecting group by treatment with 1 M HCl in methanol at
508C (100% yield). Purine 15 (61% yield)10 was prepared
from the alkylation of 6-chloro-1H-purine (16) with
potassium carbonate and 1-chloromethoxy-2-trimethylsilyl-
ethane (SEM-Cl) in DMF at 258C, which also provided
6-chloro-7-[2-(trimethylsilyl)ethoxy]methyl-1H-purine (17;
14% yield). The regiochemistry of purine 15 and its C-7
isomer 17 were determined from 2D NOESY experiments
in which their C8-H (d 8.24 and 8.36 ppm, respectively)
show correlation with their CH2N (d 5.62 and 5.80 ppm,
respectively).
Initially, identification of the structure of 6 was difficult
since it is a previously unreported compound, and reports in
the literature often do not mention N-3 alkylation under
basic conditions.8 To verify its structure, a different method
to prepare it was sought, and an adenine-tricyclic pyrone
isomer, 7, was also synthesized to compare the spectro-
scopic data and as well as the bioactivity data. A reported
method for the selective N3-alkylation of adenine9 was
followed. Treatment of mesylate 13 with adenine in N,N-
dimethylacetamide (DMA) at 1508C without base provided
compound 6 (43% yield) as the major product along with a
small amount of 5 (4% yield).
2.2. Biological activity results
ACAT mediates the esterification of intracellular choles-
terol and is believed to play a key role in lipoprotein
metabolism and atherogenesis.11 Studies of the inhibition of
ACAT with tricyclic pyrone analogs were carried out first;
the TP results are summarized in Table 1. Esters 1–3
showed marginal activity, with IC50 values of 157, 310, and
150 mM, respectively. Under the same conditions, the IC50
value of pyripyropene A was 0.1 mM. The inhibition of
CETP with these three compounds was then studied. CETP,
a hydrophobic neutral glycoprotein, mediates the transfer of
cholesteryl ester from high-density lipoprotein to low-
density lipoprotein.12 It is encouraging to find that the IC50
The synthesis of N100-adenine derivative 7 was achieved
from a palladium-mediated displacement reaction of amine
14 with chloropurine.10 Amine 14 was prepared from the
displacement of mesylate 13 with sodium azide in DMF at
608C (83% yield) followed by hydrogenation with 10% Pd/
CaCO3 in toluene under 1 atm of hydrogen at 258C (78%
yield). N-100 Adenine derivative 7 was synthesized via
palladium-mediated displacement of 6-chloro-9-[2-(tri-
methylsilyl)ethoxy]methyl-1H-purine (15)10 and amine 14
in the presence of 0.1 equiv. of Pd2(dba)3, 0.4 equiv. of
triphenylphosphine and 1.5 equiv. of potassium carbonate in

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D. H. Hua et al. / Tetrahedron 59 (2003) 4795–4803
Scheme 2.
value of compound 3 was 75 mM, while the values of
compounds 1 and 2 were .100 mM. Based on these studies,
we synthesized and evaluated a number of amides derived
from the attachment of natural amino acid at C12 of 1 and
the modification at the isopropenyl side chain such as with a
linkage of an adenine13 moiety at C14, i.e. compounds 5–7.
The N-(p-nitrophenyl)aspartamide portion of 4 was attached
in order to study whether 4 was a substrate of caspase 3.14
The in vitro study of 4 with caspase 3 showed no release of
p-nitroaniline, which indicated that 4 is not a substrate.
However, to our delight, compounds 4 and 6 (code named
CP2)5 protected from death MC65 cells that conditionally
expressed with a partial APP fusion protein (amino-17
residuesþcarboxy-99 residues15 in the absence of tetra-
cycline.5 The EC50 (effective concentration at 50%) values
of 40and 6 were 2.0 and 0.15 mM, respectively. Surprisingly,
N-9 analog 5, a regioisomer of 6, was 20-fold less active
(EC50¼3.0 mM) than 6, while N-100 analog 7, another
regioisomer of 6 was inactive. It is presently uncertain
why 6 is the most active compound, but it may be associated
with the water 0solubility of N30-adenine 6, in contrast to
that of the N9 - and N100 derivatives 5 and 7, or their
position of substitution on the adenine ring. Adenine itself
shows no activity. Since some tricyclic pyrones such as
Table 1. Biological activities of various tricyclic pyrones
Compounds
IC50, ACAT
IC50, CETP
IC50, L1210 leukemic cells
EC50, protection of APP C99-induced cell death
1
2
3
4
5
6
7
8
157 mM
310 mM
150 mM
168 mM
–
.500 mM
–
167 mM
0.1 mM
–
.100 mM
.100 mM
75 mM
.180 mM
–
.250 mM
–
–
–
–
–
–
–
–
–
–
–
30 mM
–
2.0 mM
3.0 mM
0.15 mM
.3.0 mM
.10 mM
–
–
–
.100 mM
–
Pyripyropene A
Adenine
.10 mM

D. H. Hua et al. / Tetrahedron 59 (2003) 4795–4803
4799
H10, {3-(3-pyridyl)-1H,7H-5a,6,8,9-tetrahydro-1-oxopyr-
ano-[4,3-b][1]benzopyran},16 show intense anticancer
activity, the cytotoxicity of adenine conjugated tricyclic
pyrone was examined. Compound 5 is cytotoxic to L1210
tumor cells, with IC50¼30 mM. The inability of 5 to
decrease tumor-cell viability at 1–25 mM may be ascribed
to its top pyrone A-ring, which possesses a methyl
substituent at C3 and not a 3-pyridyl substituent, which is
essential for the antitumor activity of the TP analogs.16
4.1.1. Benzyl (5aS,7S)-{7-isopropenyl-1H,7H-5a,6,8,9-
tetrahydro-1-oxopyrano[4,3-b][1]benzopyran-3-yl}ace-
tate (1). To a cold (2108C) solution of 0.68 mL (4.8 mmol)
of diisopropylamine in 20 mL of diethyl ether under argon
was added 3.00 mL (4.8 mmol; 1.6 M solution in hexanes)
of n-butyllithium via syringe, and the solution was stirred
for 1 h. In another flask, a solution of 0.50 g (1.9 mmol) of 8
in 20 mL of THF under argon was cooled to 2788C. The
LDA solution was added to the pyrone solution at 2788C
via cannula, followed by 0.67 mL (3.9 mmol) of HMPA.
After 3 h of stirring, a cold (2788C) solution of 0.55 mL
(3.9 mmol) of benzyl chloroformate in 20 mL of THF was
added via cannula, and stirred for 2 h. The reaction solution
was diluted with 40 mL of aqueous NaHCO3, and extracted
three times with dichloromethane. The combined organic
layer was washed with 40 mL of brine, dried (MgSO4),
concentrated, and column chromatographed on silica gel
using a gradient mixture of hexane and ether as eluant to
give 0.22 g (46% yield; based on reacted 8) of 1 and 0.19 g
(38% recovery) of 8. Mp 114–1168C; [a]2D3¼2288 (c 1.5,
3. Conclusions
Various tricyclic pyrones containing ester and amide side
chains at C3 were synthesized from a regioselective
deprotonation of C3 methyl group followed by carbonyl-
ation with carbon dioxide or benzyl chloroformate.
Tricyclic pyrones containing alkyl side chain at C7
possessing an adenine moiety were also synthesized by a
chemoselective hydroboration of C7 isopropenyl group
followed by functional group transformation. Chemical
modification of the functionalized tricyclic pyrones could
lead to a library of analogs for biological evaluation.
Compared with the moderate ACAT and CETP activities of
TP analogs possessing an ester side chain at C3 of the
pyrone ring (i.e. compounds 1–3), EC50 values of
compounds 4 and 6 are 2.0 and 0.15 mM, respectively, in
the protection of neuron-cell death from the toxicity of
intracellular accumulation of Ab or Ab-containing
C-terminal fragments (CTF) of APP. Thus, compounds 4
and 6 serve as lead compounds for mechanistic studies now
being carried out on intracellular Ab and CTF induced
neuron-cell death. These studies should enhance the future
development of new drugs for the prevention and treatment
of AD.
1
CHCl3); H NMR d 7.38–7.31 (m, 5H, Ar), 6.08 (s, 1H,
C10-H), 5.91 (s, 1H, C4-H), 5.28 (s, 2H, CH2O), 5.12 (dd,
J¼1.2, 5.2 Hz, 1H, C5a–H), 4.75 (s, 1H, vCH2), 4.72 (s,
1H, vCH2), 3.50 (s, 2H, CH2CO), 2.49,2.45 (m, 1H),
2.21,2.01 (m, 3H), 1.86–1.70 (m, 2H), 1.73 (s, 3H, Me),
1.34–1.25 (m, 1H); 13C NMR d 167.5, 162.6, 161.9, 156.2,
147.8, 135.2, 133.2, 128.7, 128.6, 128.4, 109.8, 109.3,
102.2, 98.8, 79.5, 67.5, 43.4, 39.9, 39.4, 32.4, 31.9, 20.8.
Anal. calcd for C24H24O5: C, 73.45; H, 6.16. Found: C,
73.74; H, 6.50.
4.1.2. Benzyl (5aS,7S)-{7-[(1R)- and (1S)-2-hydroxy-1-
methylethyl]-1H,7H-5a,6,8,9-tetrahydro-1-oxopyr-
ano[4,3-b][1]benzopyran-3-yl}acetate (2). A solution of
0.115 g (0.29 mmol) of 1 and 0.30 mL (0.30 mmol) of
BH3·THF complex (1.0 M in THF) in 5 mL of THF was
kept at 2258C for 14 h. The solution was warmed to 08C,
2 mL (0.3 mmol) of 0.5% aqueous NaOH solution and 2 mL
(17.6 mmol) of 30% hydrogen peroxide were added, and the
mixture was stirred at 08C for 6 h. The mixture was then
neutralized with 6N HCl, diluted with 30 mL of water, and
extracted three times with dichloromethane. The combined
extract was washed with brine, dried (MgSO4), concen-
trated, and column chromatographed on silica gel using a
gradient mixture of hexane and diethyl ether as eluant to
give 0.061 g (69% yield; based on reacted 1) of 2 as a
mixture of two diastereomers at C13, 2R and 2S, and
4. Experimental
4.1. General methods
Nuclear magnetic resonance spectra were obtained at
1
400 MHz for H and 100 MHz for 13C in deuteriochloro-
form, unless otherwise indicated. Mass spectra were taken
from a Hewlett–Packard 5890A Series II, GC–MS and a
Bruker Esquire 3000 Plus electrospray ionization mass
spectrometer. FAB spectra were taken by using Xe beam
(8 kV) and m-nitrobenzyl alcohol as matrix. High-resolution
Mass spectra were taken from an IonSpec HiResMALDI
mass spectrometer using 2,5-dihydroxybenzoic acid as a
matrix. (5aS,7S)-{7-Isopropenyl-3-methyl-1H,7H-5a,6,8,9-
tetrahydro-1-oxopyrano[4,3-b][1]benzopyran (8), an opti-
cally pure compound, was prepared as described.4 Pd2(dba)3
was prepared from palladium chloride, trans,trans-1,5-
diphenyl-1,4-pentadien-3-one (dibenzylidene acetone), and
sodium acetate in methanol.17 N-(t-Butyloxycarbonyl)-L-
aspartic acid 4-benzyl ester, adenine, 6-chloro-1H-purine,
and 1-(chloromethoxy)-2-(trimethylsilyl)ethane (SEM-Cl)
were purchased from Aldrich Chem. Co.
1
0.030 g (26% recovery) of 1. Mp 108–1108C; H NMR d
7.39,7.31 (m, 5H, Ar), 6.06 (s, 1H), 5.91 (s, 1H), 5.17 (s,
2H, OCH2), 5.10 (dd, J¼10.8, 5.6 Hz, 1H, C5a–H), 3.61–
3.52 (m, 2H, CH2OH), 3.50 (s, 2H, CH2CO), 2.46 (dd,
J¼14.0, 1.2 Hz, 1H), 2.13–1.96 (m, 2H), 1.73–1.11 (m,
5H), 0.91 (d, J¼2 Hz, 3H, Me); 13C NMR d 167.6, 162.7,
162.1, 156.2, 135.2, 133.8, 128.8, 128.6, 128.5, 109.0,
102.3, 98.9, 79.9, 79.8, 67.6, 65.8, 40.1, 40.0, 39.5, 39.4,
37.4, 37.3, 37.1, 32.6, 32.4, 31.2, 30.5, 28.7, 13.3 (Me for a
diastereomer), 13.2 (Me for another diastereomer). Anal.
calcd for C24H26O6·0.5H2O: C, 68.65; H, 6.49. Found: C,
69.22; H, 6.54.
4.1.3. Benzyl (5aS,7S,9aS,10S) -{10-hydroxy-7-[(1R) and
(1S)-2-hydroxy-1-methylethyl]-1H,7H-5a,6,8,9-tetra-
hydro-1-oxopyrano[4,3-b][1]benzopyran-3-yl}acetate
The reported procedures for the inhibitory activities of
ACAT,18 CETP,19 L1210 tumor cells,16 and APP C99-
induced cell death5 were followed.

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D. H. Hua et al. / Tetrahedron 59 (2003) 4795–4803
(3). A solution of 0.10 g (0.26 mmol) of 1 and 0.26 mL
(0.26 mmol) of BH3·THF complex (1.0 M in THF) in 3 mL
of THF was kept at 2208C for 3 h and then at 08C for 12 h.
To it, 2 mL of 0.5% aqueous NaOH solution and 2 mL of
30% hydrogen peroxide were added, and the mixture was
stirred at 08C for 4 h. The mixture was then neutralized with
6N HCl, diluted with 10 mL of water, and extracted three
times with dichloromethane. The combined extract was
washed with brine, dried (MgSO4), concentrated, and
column chromatographed on silica gel using a gradient
mixture of hexane and diethyl ether as eluant to give 15 mg
(14% yield) of 2 and 36 mg (32% yield) of 3. 1H NMR 7.36
(m, 5H, Ar), 5.99 (s, C4H), 5.18 (s, CH2O), 4.66 (dd, J¼8.8,
3.2 Hz, 1H, C10H), 4.46 (m, 1H, C5aH), 4.26 (bs, 1H, OH),
3.58 (m, 2H, CH2O), 3.53 (s, 2H, CH2CO), 2.21 (m, 1H),
2.14 (bs, 1H, OH), 1.78–1.33 (a serious of m, 8H), 0.93 (d,
J¼7 Hz, 3H, Me of a diastereomer), 0.90 (d, J¼7 Hz, 3H,
Me of another diasteromer). 13C NMR (2 diastereomers at
C13) d 167.3, 164.6, 163.4, 156.6, 135.0, 128.7 (2C, Ar),
128.4 (2C, Ar), 103.0, 100.9, 94.0, 67.5, 65.9, 59.5, 39.5,
39.1, 37.9, 31.6, 29.6, 24.0, 22.3, 13.7, 13.5. Anal. calcd for
C24H28O7·0.5H2O: C, 65.89; H, 6.68. Found: C, 66.17; H,
7.26.
benzyl ester protected amide. A solution of 40 mg
(0.06 mmol) of the above amide and 1 mL of 1% aqueous
NaOH in 2 mL of THF was stirred at 258C for 1 h. The
solution was neutralized with 4N HCl, concentrated, and the
resulting solid was washed with dichloromethane (to
remove benzyl alcohol) and dried under vacuum to give
34 mg (100% yield) of 4. Mp 234–2368C; [a]2D3¼þ23.18 (c
1.2, CHCl3); MS (electrospray), m/z 537 (Mþ), 535 (M22),
1
518 (100%, M21–H2O). H NMR (DMSO-d6) d 8.19 (d,
J¼9.6 Hz, 1H, Ar), 7.85 (d, J¼9.6 Hz, 1H, Ar), 6.11 (s, 1H,
C4H), 5.94 (s, 1H, C 10H), 4.75 (s, 1H, vCH2), 4.72 (s, 1H,
vCH2), 4.05 (m, 1H, CHN), 3.24 (s, 2H, CH2CO), 2.10 (m,
2H, CH2CO), 1.90–1.02 (a series of m, 7H), 1.70 (s, 3H,
Me); 13C NMR (DMSO-d6) d 173.9, 170.8, 169.8, 165.4,
162.0, 157.0, 148.1, 145.6, 141.9, 133.1, 128.2, 124.9,
118.8, 109.9, 100.6, 90.3, 67.9, 55.8, 49.9, 34.7, 31.5, 29.0,
25.4, 20.6, 17.2. HRMS calcd for C27H27N3O9 537.1748,
found 537.1742.
4.1.6. Benzyl (S)-3-amino-4-oxo-4-(4-nitrophenylamino)-
butanoate (10). To a solution of 0.214 g (1.55 mmol) of
p-nitroaniline and 0.50 g (1.55 mmol) of N-(t-butyloxy-
carbonyl)-L-aspartic acid 4-benzyl ester (11) in 10 mL of
dichloromethane at 258C under argon, was added a solution
of 0.383 g (1.86 mmol) of DCC in 20 mL of dichloro-
methane via cannula. The reaction mixture was stirred for
24 h, filtered, concentrated, and column chromatographed
on silica gel using a mixture of hexane, methanol, and
dichloromethane (40:4:1) as eluant to give 0.45 g of the
amide (85% yield; based on reacted p-nitroaniline) and
0.05 g (23% recovery) of p-nitroaniline. Mp 47–508C;
[a]2D0¼236.58 (c 1.3, CHCl3); 1H NMR d 9.18 (s, 1H, NH),
8.20 (d, J¼7 Hz, 2H, Ar), 7.66 (d, J¼7 Hz, 2H, Ar), 7.36 (s,
5H, Ar), 5.86 (s, 1H, NH), 5.20 (d, J¼12.4 Hz, 1H, OCH2),
5.16 (d, J¼12.4 Hz, 1H, OCH2), 4.67 (m, 1H, CHN), 3.09
(dd, J¼17, 4 Hz, 1H, CH2CO), 2.82 (dd, J¼17, 6 Hz, 1H,
CH2CO), 1.49 (s, 9H, t-Bu); 13C NMR d, 171.6, 169.4,
153.1, 143.4, 135.2, 128.6, 128.5, 128.47, 128.2, 125.0,
119.2, 67.1, 65.8, 51.6, 35.4, 28.2. A solution of 0.15 g
(0.34 mmol) of the above amide and 0.85 mL (3.39 mmol)
of 4 M HCl in dioxane was stirred at 258C for 2 h,
neutralized with 10 mL of aqueous NaHCO3, and extracted
three times with dichloromethane. The combined organic
layer was washed with brine, dried (MgSO4), and
concentrated to give 0.122 g of 10 (100% yield):
[a]2D2¼2198 (c 0.15, CHCl3; 1H NMR d 10.00 (s, 1H,
CONH), 8.22 (d, J¼7 Hz, 2H, Ar), 7.75 (d, J¼7 Hz, 2H,
Ar), 7.34 (s, 5H, Ar), 5.18 (d, J¼12.4 Hz 1H, OCH2), 5.14
(d, J¼12.4 Hz, 1H, OCH2), 3.83 (dd, J¼6.8, 4.4 Hz, 1H,
CHN), 3.02 (dd, J¼17, 4.4 Hz, 1H, CH2), 2.96 (dd, J¼17,
6.8 Hz, 1H, CH2); 13C NMR d, 174.4, 172.0, 143.4, 135.1,
128.7, 128.5, 128.4, 128.3, 125.1, 118.9, 66.9, 52.4, 38.9.
HRMS calcd for C17H18N3O5 (Mþ1) 344.1169, found
344.1150.
4.1.4. (5aS,7S)-{7-Isopropenyl-1H,7H-5a,6,8,9-tetra-
hydro-1-oxopyrano[4,3-b][1]-benzopyran-3-yl}acetic
acid (9). To a cold (2108C) solution of 0.27 mL
(1.90 mmol) of diisopropylamine in 5 mL of diethyl ether
under argon was added 1.20 mL (1.90 mmol; 1.6 M solution
in hexanes) of n-butyllithium via syringe, and the solution
was stirred for 1 h. In another flask, a solution of 0.25 g
(0.97 mmol) of 8 in 5 mL of THF under argon was cooled to
2788C. To it, the LDA solution was added via cannula,
stirred for 2 h, and carbon dioxide gas was then introduced.
The color of the blue anion changed to brownish color. The
reaction mixture was stirred for 30 min, 20 mL of aqueous
NaHCO3 was added, and extracted with diethyl ether three
times. The aqueous layer was acidified with 6N HCl,
extracted three times with dichloromethane. The combined
dichloromethane layers were washed with water, brine,
dried (MgSO4), and concentrated to give 0.271 g (92%
yield) of 9. This material was used in the next step without
further purification. Compound 9 undergoes decarboxyl-
ation when subjected to silica gel column chromatography
to give 8. [a]2D3¼þ228 (c 1.0, CHCl3); MS, EI m/z 258
(M2CO2), 189, 176; 1H NMR d 6.09 (s, 1H), 5.95 (s, 1H),
5.13 (m, 1H), 4.75 (s, 1H, vCH2), 4.72 (s, 1H, vCH2), 3.51
(s, 2H, CH2CO), 2.51–1.21 (a series of m, 7H), 1.73 (s, 3H,
Me); 13C NMR d 171.0, 163.2, 162.8, 156.5, 147.8, 133.6,
110.0, 109.2, 102.6, 98.9, 79.8, 43.5, 40.0, 39.3, 32.6, 32.1,
20.9. HRMS calcd for C17H19O5 (Mþ1) 303.1457, found
303.1400.
4.1.5. (3S)-3-{[(5a0S,70S)-7-Isopropenyl-1H,7H-5a,6,8,9-
tetrahydro-1-oxopyran[4,3-b][1]benzopyran-3-
yl]methylcarbamoyl}-4-oxo-4-(4-nitrophenylamino)-
butanoic acid (4). A mixture of 35 mg (0.12 mmol) of 9,
44 mg (0.12 mmol) of 10, and 36 mg (0.18 mmol) of DCC
in 5 mL of dichloromethane under argon was stirred at 258C
for 20 h. The reaction mixture was filtered, and the filtrate
was concentrated, and column chromatographed on silica
gel using a gradient mixture of hexane, dichloromethane
and methanol as eluant to give 72 mg (90% yield) of the
4.1.7. (5aS,7S)-7-[(1R) and (1S)-2-Hydroxy-1-methyl-
ethyl]-3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-oxopyr-
ano[4,3-b][1]benzopyran (12). To a cold solution (2258C)
of 0.50 g (1.94 mmol) of 8 in 10 mL of THF under argon,
was added 1.94 mL (1.94 mmol) of BH3·THF complex
(1.0 M in THF). After stirring the solution at 2258C for
10 h, 14 mL (2.1 mmol) of 0.5% aqueous NaOH and 4 mL
of 30% hydrogen peroxide were added at 08C. The solution

D. H. Hua et al. / Tetrahedron 59 (2003) 4795–4803
4801
was stirred for 4 h, diluted with 50 mL of water, and
extracted three times with dichloromethane. The combined
organic layer was washed with 40 mL of brine, dried
(MgSO4), concentrated, and column chromatographed on
silica gel using a gradient mixture of hexane and diethyl
ether as eluants to give 0.326 g (82% yield; based on reacted
8) of 12 as a mixture of two diastereomers at C12 (1:1; based
on 13C NMR spectrum) and 0.130 g (26% recovery) of 8. 1H
NMR d 6.08 (s, 1H, C4H), 5.71 (s, 1H, C10H), 5.07 (t,
J¼5.2 Hz, 1H, C5aH), 3.62–3.52 (m, 2H, CH2O), 2.46 (m,
1H), 2.19 (s, 3H, Me), 2.13–1.99 (m, 2H), 1.73–1.51 (m,
3H), 1.19–1.12 (m, 2H), 0.92 (d, J¼7 Hz, 3H, Me); 13C
NMR (two diastereomers) d 163.5, 162.8, 161.6, 133.0,
109.0, 100.0, 97.4, 79.7, 79.6, 65.6, 39.9, 39.8, 39.4, 37.2,
37.1, 36.9, 32.4, 32.3, 31.1, 30.4, 28.5, 20.1, 13.2 (Me for a
diastereomer), 13.1 (Me for another diastereomer). Anal.
calcd for C16H20O4: C, 69.55; H, 7.29. Found: C, 69.26; H,
7.03.
5 (less polar; two diastereomers at C12): mp 228–2308C;
MS, electrospray, m/z 394 (Mþ1; 100%), 259 (M2
0
1
adenine), 136 (adenine); H NMR d 8.36 (s, 1H, C2 H),
7.78 (s, 1H, C80H), 6.09 (s, 1H, C4H), 5.89 (bs, 2H, NH2;
disappeared when 1 drop of D2O was added), 5.72 (s, 1H,
C10H), 5.01 (m, 1H, C5aH), 4.24 (dd, J¼14, 7 Hz, 1H,
CHN), 4.01 (dd, J¼14, 7 Hz, 1H, CHN), 2.5–1.2 (a serious
of m, 8H), 2.19 (s, 3H, Me), 0.90 (d, J¼7 Hz, 3H, Me); 13C
NMR d 163.4, 162.7, 161.9, 155.6 (adenine moiety), 153.5
(adenine moiety), 150.6 (adenine moiety), 140.9 (adenine
moiety), 132.1, 119.8 (adenine moiety), 109.9, 99.9, 97.5,
79.4, 79.2, 47.9, 39.3, 38.4, 38.3, 38.1, 38.0, 36.2, 32.3,
32.1, 31.1, 27.8, 20.3, 13.8. Anal. calcd for C21H23N5O3: C,
64.11; H, 5.89. Found: C, 63.80; H, 5.93.
Compound 6 (more polar; 2 diastereomers at C12): MS,
electrospray, m/z 394 (Mþ1, 100%), 259 (M2adenine), 136
0
1
(adenine); H NMR d 8.07 (s, C8 H of adenine), 7.98 and
7.97 (2 s, 1H, C20H of adenine; 2 diastereomers), 6.10 (s,
1H, C10H), 5.72 and 5.71 (2s, 1H, C4H), 5.02 (m, 1H,
C5aH), 4.50 (dd, J¼14, 7 Hz, 1H, CHN), 4.08 (2dd, J¼14,
8 Hz, 1H, CHN; 2 diastereomers), 2.46 (m, 2H), 2.20 and
2.19 (2s, 3H, Me; 2 diastereomers), 2.10–1.22 (a series of
m, 6H), 0.91 (d, J¼7.0 Hz, 3H, Me). 13C NMR (2
diastereomers) d 163.2 and 163.1, 162.4, 161.7, 154.4,
154.0, 150.7, 142.3, 131.7 and 131.6, 121.0, 199.8, 99.7,
97.3, 79.0, 78.8, 54.5 and 54.4, 38.9, 38.1 and 38.0, 37.1 and
36.9, 36.1, 32.0 and 31.9, 30.7, 27.6, 20.1, 13.3 and 13.2.
HRMS calcd for C21H24N5O3 (MþH) 394.1881, found
394.1875. Anal. calcd for C21H23N5O3·2H2O: C, 58.73; H,
6.34. Found: C, 59.20; H, 6.18.
4.1.8. (5aS,7S)-3-Methyl-7-[(1R) and (1S)- 2-(methane-
sulfonyloxy)-1-methylethyl]-1H,7H-5a,6,8,9-tetrahydro-
1-oxopyrano[4,3-b][1]benzopyran (13). To a cold (08C)
solution of 50 mg (0.18 mmol) of 12 in 5 mL of dichloro-
methane under argon, were added 0.08 mL (0.54 mmol) of
triethylamine and 0.02 mL (0.27 mmol) of methanesulfonyl
chloride. The solution was stirred for 3 h, diluted with
30 mL of water, and extracted three times with dichloro-
methane. The combined dichloromethane layer was washed
with saturated aqueous NaHCO3, and brine, dried (MgSO4),
concentrated, and column chromatographed on silica gel
using a gradient mixture of hexane and ether as eluants to
give 60 mg (94% yield) of 13 as a mixture of two
diastereomers (1:1; based on 13C NMR spectrum). 1H
NMR d 6.08 (s, 1H, C4H), 5.71 (s, 1H, C10H), 5.06 (m, 1H,
CHO), 4.18–4.08 (m, 2H, CH2O), 3.03 (s, 3H, MeS), 2.49
(d, J¼2.8 Hz, 1H), 2.19 (s, 3H, Me), 2.14–1.11 (m, 7H),
0.98 (d, J¼6.8 Hz, 3H, Me); 13C NMR d 163.2, 162.4,
161.7, 132.1, 109.6, 105.2, 99.8, 79.2, 79.1, 72.3, 38.9, 37.5,
37.4, 37.3, 37.2, 36.9, 32.2, 32.1, 30.8, 28.6, 20.2, 13.3 (Me
for a diastereomer), 13.2 (Me for another diastereomer).
Anal. calcd for C17H22O6S: C, 57.61; H, 6.26. Found: C,
57.65; H, 6.43.
4.1.10. Synthesis of compound 6 from adenine in DMA.
A solution of 0.21 g (0.59 mmol) of mesylate 13 and 80 mg
(0.59 mmol) of adenine in 3 mL of DMA (freshly distilled
from CaCl2 under reduced pressure) was heated at 1508C for
3 h. DMA was removed under reduced pressure (708C/
0.5 mm Hg), and the residue of the distillation was triturated
with 5 mL of dichloromethane. To the residue, 50 mg
(0.59 mmol) of NaHCO3 and 3 mL of ethanol were added,
and the mixture was subjected to a silica gel column
chromatography using a gradient mixture of dichloro-
methane and ethanol as eluant to give 0.10 g (43% yield)
of 6 and 0.01 g (4% yield) of 5.
4.1.9. (5aS,7S)-7-[(1R) and (1S)-2-(N9-Adenyl)-1-methyl-
ethyl]-3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-oxopyr-
ano[4,3-b][1]benzopyran (5) and (5aS,7S)-7-[(1R) and
(1S)- 2-(N3-adenyl)-1-methylethyl]-3-methyl-1H,7H-
5a,6,8,9-tetrahydro-1-oxopyrano[4,3-b][1]benzopyran
(6). To a solution of 0.008 g (0.34 mmol) of NaH in 3 mL of
DMF under argon was added 0.046 g (0.34 mmol) of
adenine at 258C, and the solution was stirred for 1 h. This
solution was then added to a solution of 0.110 g
(0.30 mmol) of 13 in 3 mL of DMF via cannula. The
reaction solution was stirred at 808C for 20 h. The solvent,
DMF, was removed under vacuum at 508C, 10 mL of
dichloromethane was added to the residue, and the
dichloromethane was removed via a pipette (to remove
trace of DMF and tricyclic pyrone by-product 12). To the
resulting solids, 20 mL of ethanol was added, filtered, and
the filtrate was concentrated, and column chromatographed
on silica gel using a gradient mixture of dichloromethane
and methanol as eluant to give 0.026 g (21% yield) of 5 (less
polar) and 0.006 g (5% yield) of 6 (more polar). Compound
4.1.11. (5aS,7S)-7-[(1R) and (1S)-2-Azido-1-methyl-
ethyl]-3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-oxopyr-
ano[4,3-b][1]benzopyran (18). A solution of 0.70 g
(2.0 mmol) of mesylate 13 and 0.26 g (4.0 mmol) of sodium
azide in 20 mL of DMF under argon was stirred at 608C for
16 h. The solution was cooled, diluted with ether, washed
twice with water, and brine, dried (MgSO4), concentrated,
and column chromatographed on silica gel using a gradient
mixture of hexane and ethyl acetate to give 0.50 g (83%
yield) of 18 and 60 mg (9% recovery) of 13. Compound 18
(2 diastereomers at C12): mp 87–888C; MS, m/z 301 (Mþ);
1H NMR d 6.09 (s, 1H, C4H), 5.71 (s, 1H, C10H), 5.10 (m,
1H, C5aH), 3.30 (dd, J¼13, 6 Hz, 1H, CHN), 3.23 (dd,
J¼13, 7 Hz, 1H, CHN), 2.48 (d, J¼14 Hz, 1H), 2.19 (s, 3H,
Me), 2.10–1.95 (m, 2H), 1.7–1.5 (m, 4H), 1.27–1.08 (m,
1H), 0.95 (d, J¼7 Hz, 3H, Me); 13C NMR d 163.4, 162.7,
161.8, 132.4, 109.6, 99.9, 97.5, 79.5 and 79.4 (C5a, 2
isomers), 55.5, 39.2, 38.2 and 38.15 (2 isomers), 37.8 and

4802
D. H. Hua et al. / Tetrahedron 59 (2003) 4795–4803
37.78 (2 isomers), 37.0, 32.3 and 32.2 (2 isomers), 30.9,
28.6, 20.3, 14.5. Anal. calcd for C16H19N3O3: C, 63.77; H,
6.35. Found: C, 63.99; H, 6.51.
NMR d 8.40 (s, 1H), 7.89 (s, 1H), 6.07 (s, 1H, C4H), 5.81
(bs, 1H, NH), 5.70 (s, 1H, C10H), 5.56 (s, 2H, NCH2O),
5.03 (m, 1H, C5aH), 3.64 (m, 1H, CHN), 3.60 (t, J¼8 Hz,
2H, CH2O), 3.50 (m, 1H, CHN), 2.46 (d, J¼12 Hz, 1H),
2.18 (s, 3H, Me), 2.10–1.2 (a serious of m, 7H), 0.93 (d,
J¼7 Hz, 3H, Me; 2 diastereomers as indicated by 2 sets of
doublet), 0.92 (t, J¼8 Hz, 2H, CH2Si, 2 diastereomers as
indicated by 2 sets of triplet), 20.05 (s, 9H, MeSi); 13C
NMR d 163.5, 162.7, 161.8, 155.3, 153.8, 140.1, 132.6,
109.5, 100.0, 97.6, 79.7 and 79.5 (C5a; 2 diastereomers),
72.2, 67.4, 39.5, 38.4 and 38.3 (2 diastereomers), 38.1, 36.7,
32.5 and 32.3 (2 diastereomers), 31.3, 28.3, 20.3, 17.9, 14.3
and 14.2 (2 diastereomers), 21.2. HRMS calcd for
C27H38N5O4Siþ (MþHþ) 524.2695, found 524.2292.
4.1.12. (5aS,7S)-7-[(1R) and (1S)-2-Amino-1-methyl-
ethyl)-3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-oxopyr-
ano[4,3-b][1]benzopyran (14). A solution of 0.42 g
(1.4 mmol) of azide 18 and 1.0 g of 10% Pd/CaCO3 in
toluene was stirred under 1 atm. of hydrogen at 258C for
14 h. The mixture was filtered through a short silica gel
column using ethyl acetate, CHCl3/CH3OH (2:1), and then
CHCl3/CH3OH/NH4OH (2:1:0.01) as eluants to give 0.30 g
(78% yield) of amine 14 (2 diastereomers at C12). 1H NMR
d 6.07 (s, 1H, C4H), 5.71 (s, 1H, C10H), 5.07 (m, 1H,
C5aH), 2.73 (m, 1H, 2 isomer, CHN), 2.58 (m, 1H, 2
isomers, CHN), 2.46 (d, J¼13 Hz, 1H), 2.19 (s, 3H, Me),
2.15–1.10 (m, 7H), 0.91 (d, J¼7 Hz, 3H, Me); 13C NMR d
163.4, 162.7, 161.7, 132.9, 109.3, 100.0, 97.5, 79.8 and 79.7
(2 isomers), 66.0, 46.0 and 45.9 (2 isomers), 40.9, 39.1, 38.5
and 38.4 (2 isomers), 37.1, 32.6 and 32.4 (2 isomers), 31.3,
31.2, 28.6, 20.3, 15.5, 14.3. HRMS calcd for C17H22NO3
(Mþ1) 276.1601, found 276.1610.
4.1.15. (5aS,7S)- 7-[(1R) and (1S)-2-(N10-Adenyl)-1-
methylethyl]-3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-
oxopyrano[4,3-b][1]benzopyran (7). A solution of 4 mg
(0.008 mmol) of SEM-adenine 19 in 1 mL each of 1 N HCl
and methanol was heated at 508C for 6 h and cooled to 258C.
The solution was neutralized with saturated aqueous
NaHCO3, extracted with ethyl acetate, and the organic
layer was washed with brine, dried (MgSO4), concentrated,
and column chromatographed on silica gel column using a
mixture of CHCl3/MeOH (10:1) as eluant to give 3.0 mg
(97% yield) of 7. 1H NMR d 8.44 (s, 1H), 8.00 (s, 1H), 6.3
(bs, 1H, NH), 6.08 (s, 1H, C4H), 5.72 (s, 1H, C10H), 5.05
(m, 1H, C5aH), 3.75 (m, 1H, CHN), 3.57 (m, 1H, CHN),
2.47 (d, J¼14 Hz, 1H), 2.19 (s, 3H, Me), 2.10–1.2 (a serious
of m, 7H), 1.01 (d, J¼7 Hz, 3H, Me); 13C NMR d 163.5,
162.8, 161.8, 152.8, 138.3 (2 C), 133.7, 132.6, 131.6, 109.5,
100.0, 97.6, 79.7 and 79.5 (2 isomers at C12), 39.5, 38.5,
38.3, 38.1, 36.7, 32.3, 31.3, 29.9, 28.4, 20.3, 14.3 and 14.2
(2 isomers). Anal. calcd for C21H23N5O3: C, 64.11; H, 5.89.
Found: C, 64.35; H, 6.10.
4.1.13. 6-Chloro-9-[(2-trimethylsilylethoxy)methyl]-1H-
purine (15).15 A solution of 0.31 g (2.0 mmol) of 6-chloro-
1H-purine and 0.83 g (6.0 mmol) of potassium carbonate in
15 mL of DMF was stirred at 258C for 20 min. under argon.
To it, 0.53 mL (3.0 mmol) of 1-(chloromethoxy)-2-(tri-
methylsilyl)ethane (SEM-Cl) was added via syringe, the
solution was stirred for 9 h, filtered through Celite, and the
filtrate was diluted with diethyl ether, washed with water,
and brine, dried (MgSO4), concentrated, and column
chromatographed on silica gel using a gradient mixture of
hexane and ethyl acetate to give 0.35 g (61% yield) of 15
and 0.08 g (14% yield) of 6-chloro-3-[(2-trimethylsilyl-
ethoxy)methyl]-1H-purine (17). Compound 15: mp 33–
1
348C; H NMR d 8.70 (s, 1H), 8.24 (s, 1H), 5.62 (s, 2H,
NCH2O), 3.60 (t, J¼8 Hz, 2H, CH2O), 0.94 (t, J¼8 Hz, 2H,
CH2Si), 20.12 (s, 9H, MeSi); 13C NMR d 152.4, 152.2,
151.3, 145.4, 131.5, 72.7 (CN), 67.8 (CO), 17.8, 21.4 (CSi).
Acknowledgements
We thank Drs Charles E. Chandler and Yvette Savoy,
Department of Cardiovascular and Metabolic Diseases,
Pfizer Global Research and Development, Gronton Labora-
tories, Pfizer Inc., Eastern Point Road, Groton, CT 06340,
for carrying out a part of the ACAT inhibition studies. D. H.
H. gratefully acknowledges financial support from the
National Science Foundation (CHE-0078921), National
Institutes of Health (National Cancer Institute, CA86842),
American Heart Association, Heartland Affiliate
(0051658Z, 9951177Z), NASA-BioServe Space Tech-
nologies (NAGW-1197), and Great Plains Diabetes
Research, Inc.
6-Chloro-7-[(2-trimethylsilylethoxy)methyl]-1H-purine
1
(17). Mp 80–818C; H NMR d 8.93 (s, 1H), 8.36 (s, 1H),
5.80 (s, 2H, NCH2O), 3.61 (t, J¼8 Hz, 2H, CH2O), 0.94 (t,
J¼8 Hz, 2H, CH2Si), 20.02 (s, 9H, MeSi); 13C NMR d
162.5, 153.0, 149.3, 143.7, 122.5, 75.9 (CN), 67.2 (CO),
17.9, 21.3 (CSi). Anal. calcd for C11H17ClN4OSi: C, 46.39;
H, 6.02. Found: C, 46.52; H, 6.08.
4.1.14. (5aS,7S)-3-Methyl-7-{(1R) and (1S)-2-[(N9-tri-
methylsilylethoxymethyl)-N10-adenyl]-1-methylethyl}-
1H,7H-5a,6,8,9-tetrahydro-1-oxopyrano[4,3-b][1]benzo-
pyran (19). A mixture of 56 mg (0.20 mmol) of amine 14,
114 mg (0.40 mmol) of purine 15, 18 mg (0.02 mmol) of
Pd2(dba)3, 21 mg (0.08 mmol) of Ph3P, and 41 mg
(0.30 mmol) of K2CO3 in 5 mL of toluene was heated in a
sealed tube at 1408C. After stirring for 5 h, the mixture was
cooled to 258C, diluted with water, and extracted three times
with diethyl ether. The combined ether layer was washed
with water, and brine, dried (MgSO4), concentrated, and
column chromatographed on silica gel using a gradient
mixture of dichloromethane and ethyl acetate as eluant to
give 17 mg (16% yield) of 19 (2 diastereomers at C12). 1H
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