Synthesis and HIV inhibition activity of 2',3'-dideoxy-3'-C- hydroxymethyl nucleosides

Article abstract of DOI:10.1021/jm950822k

A series if 2',3'-dideoxy-3'-C-hydroxymethyl purine nucleosides were prepared based on the photochemical ring expansion of a chiral cyclobutanone precursor, (2S)-trans-2,3-bis[(benzoyl-oxy)methyl]cyclobutanone, in the presence of a 6-substituted purine. B

Full text of DOI:10.1021/jm950822k

5
276
J . Med. Chem. 1996, 39, 5276-5280
Notes
Syn th esis a n d HIV In h ibition Activity of 2′,3′-Did eoxy-3′-C-h yd r oxym eth yl
Nu cleosid es
E. Lee-Ruff,*, Mario Ostrowski, Azim Ladha,1b Dennis V. Stynes,1a Isaak Vernik,1a J i-Long J iang,1a
1a
1b
1
c
1b
,1b
Wei-Qin Wan, Shi-Fa Ding, and Sadhna J oshi*
Department of Chemistry, York University, Toronto, Ontario M3J 1P3, Canada, and Department of Microbiology, University of
Toronto, Toronto, Ontario M5S 1A8, Canada
X
Received November 7, 1995
A series if 2′,3′-dideoxy-3′-C-hydroxymethyl purine nucleosides were prepared based on the
photochemical ring expansion of a chiral cyclobutanone precursor, (2S)-trans-2,3-bis[(benzoyl-
oxy)methyl]cyclobutanone, in the presence of a 6-substituted purine. Both R- and â-anomers
are produced in this transformation. Deprotection was effected by reaction of the photoadducts
with saturated methanolic ammonia. Nine purine nucleosides were tested for their inhibitory
effect of HIV IIIB virus on H9 cells. The 6-hexyloxy and adenine derivatives 4e,c, respectively,
appeared to be most effective at inhibiting viral reproduction with 4c comparable in activity
to ddI and AZT.
In tr od u ction
Structurally modified nucleosides have become rec-
2
3
ognized as potential antiviral and anticancer agents.
Specifically, certain 2′,3′-dideoxy ribonucleosides have
been shown to protect ATH8 and MT4 cells against the
cytopathology of the human immunodeficiency virus
(
HIV-1), the causative agent of acquired immune defi-
4
ciency syndrome (AIDS). These nucleosides include
-amino-, 6-halo-, and 6-alkoxypurine-substituted de-
6
rivatives. We have recently reported on a new method
7
for the synthesis of 2′,3′-dideoxynucleosides and 2′,3′-
dideoxy-3′-C-hydroxymethyl nucleosides5 based on the
photochemical ring expansion of chiral cyclobutanones.
Our method generates both anomers of these chiral
nucleosides. In view of the activity reported for 6-alkoxy-
,6
F igu r e 1. Crystal structure of nucleoside 4c.
on HIV-1 replication. Their effect was compared with
that of AZT and ddI.
4
purine 2′,3′-dideoxy nucleosides as HIV inhibitors
(specifically the 6-(hexyloxy)purine derivatives) and the
activity of some 2′,3′-dideoxy-3′-C-(hydroxymethyl)ribo-
8
Syn th esis
sidic nucleosides, we were interested in extending our
novel synthesis to include some other 6-substituted
purine 2′,3′-dideoxy-3′-C-hydroxymethyl nucleosides and
to test their HIV inhibitory effect. These latter com-
pounds are close structural analogues to the natural 2′-
deoxy nucleosides and the naturally occurring oxetano-
cin along with their derivatives which have been shown
The chiral cyclobutanone 1 was prepared according
6
13
to a modified procedure as described by Ahmad. The
optical purity of this ketone was in excess of 98%.6
Irradiation of ketone 1 in the presence of alcohols or
reagents incorporating acidic N-H functions gives rise
to ring expansion products and cycloelimination as the
principal pathways. For example, irradiation of this
ketone in the presence of 6-chloropurine resulted in the
formation of an anomeric mixture of the protected
nucleoside 2b and the cycloelimination product 3 in 43%
and 10% yields, respectively. Photoadducts 2a ,b were
reported in our recent study of the photochemical ring
expansion of ketone 1 in the presence of purine and
9
to be effective as anti-HIV-1 and anti-herpes virus
1
0
agents. Two other known nucleoside analogues, azi-
dothymidine (zidovudine, AZT) and dideoxyinosine
(didanosine, ddI), have been found to inhibit viral
replication and to improve survival and reduce infection
1
1,12
rate in patients with AIDS.
on the synthesis of the R- and â-anomers of 6-chloro-,
-amino-, 6-methoxy-, 6-hexyloxy-, and the unsubsti-
In this study we report
6
6
6-chloropurine, respectively. Irradiation of ketone 1 in
tuted purin-9-yl-2′,3′-dideoxy-3′-C-(hydroxymethyl)ri-
bosides from the photochemical ring expansion of chiral
ketone 1, as well as the HIV-1 inhibition studies of
these. Two of these compounds showed inhibitory effect
acetonitrile in the presence of 6-methoxypurine or
6-(hexyloxy)purine gave the corresponding photoadducts
2d ,e in 41% and 30% yields, respectively, along with
some cycloelimination product 3 (15%).
The photoadduct 2b could be converted to the adenine
derivatives 4c and 5c with methanolic ammonia at 100
X
Abstract published in Advance ACS Abstracts, November 15, 1996.
S0022-2623(95)00822-3 CCC: $12.00 © 1996 American Chemical Society

Notes
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 26 5277
Sch em e 1
°
C in a sealed tube. Separation of 4c and 5c could be
achieved by preparative HPLC. A crystal structure of
c was determined (see Figure 1) and confirmed the
4
assignment of the regiochemistry for the oxacarbene
insertion to the N(9)-H position of the purine ring in
the original photoadduct 2b. The crystal structure also
confirms the absolute structure assignment of 4c as the
â-anomer. Under the conditions for debenzoylation
employed (100 °C in sealed tube), ammonia acts as an
efficient nucleophile to displace the chloro group. How-
ever, when debenzoylation was effected in methanolic
ammonia under ambient conditions, no adenine nucleo-
side 4c or 5c was produced. Instead anomeric mixtures
of the chloro nucleosides 4b and 5b and methoxy
nucleosides 4d and 5d were formed in 19% and 41%
yields, respectively. Under the latter conditions, metha-
nol acts as a nucleophile to displace the chloride
substituent.
Debenzoylation of the photoadducts 2a ,d ,e gave an
anomeric mixture of the corresponding deprotected
nucleosides 4 and 5 in good yields. The anomeric
mixture could be separated by preparative thin-layer
chromatography. Assignment of the R- and â-anomeric
F igu r e 2. H9 cell infection by HIV-1 IIIB in the presence of
5
0 µM adenosine analogues: (light gray) day 7 and (dark gray)
day 11.
1
associated with H-1′ and H-4′, whereas no such correla-
tion could be observed for the R-anomers 5.
configurations was based on characteristic H NMR
features in the 1-D and 2-D COSY and NOESY spectra.
It has been shown that ring protons syn to the base are
In h ibition Stu d ies
more deshielded than those which are anti.1
4-16
The
anomer which has a downfield shift for H-4′ was
assigned to the R-anomer. In each of the R-anomers 5,
the H-4′ signal was observed at 4.1 ppm, whereas that
for the â-anomers 4 appeared at 4.3 ppm. This aniso-
tropic effect of the base was also apparent in the
chemical shifts of both H-2′ and -2′′ protons where the
signals for the R-anomers were well resolved whereas
the peaks for the â-anomers were overlapped. The
former showed three separate multiplets for H-2′, H-2′′,
and H-3′, whereas the latter showed only two multiplets.
Further confirmation was obtained from 2-D NOESY
spectroscopy of the two anomeric series 4 and 5. The
â-anomers 4 exhibited a NOE correlation for the signals
The effect of the nine purine nucleosides on HIV-1
replication was examined in H9 cells. All nine com-
pounds showed no evidence of cytotoxity to H9 cells
while in culture over 14 days at the concentrations
tested (1-100 µM; data not shown). Of the nine
derivatives tested, only compounds 4c,e significantly
inhibited acute infection of H9 cells by the laboratory
strain HIV IIIB (Figure 2). Although compounds 4b
and 5b (anomeric mixture) inhibited virus infection at
day 7, the effect was lost by day 11 in the culture (Figure
2). The antiviral effect of compounds 4c,e was dose
dependent with the higher concentration (50 µM) being
more effective at inhibiting virus at day 14 (Figure 3).

5
278 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 26
Notes
F igu r e 5. Day of infection syncytia occurred at 50 µM
nucleoside (*tested as an anomeric mixture).
F igu r e 3. Results of H9 cell infection by HIV-1 IIIB in the
presence of compounds 5a and 4e,c: (black) day 7 and (gray)
day 14.
F igu r e 6. Results of H9 cell infection by HIV-1 IIIB in the
presence of compounds 4c,e, ddI, and AZT: (black) day 9 and
(gray) day 15.
clinical isolates of HIV-1 as well as the antiviral effects
against AZT and ddI resistant isolates of HIV-1.
Exp er im en ta l Section
Gen er a l. Melting points (mp) were determined on a
Reichert melting point apparatus and are uncorrected. Ul-
traviolet (UV) absorption spectra were recorded on an HP8452a
diode array spectrophotometer. Infrared (IR) spectra were
obtained on a Perkin Elmer 1310 spectrometer as thin films
or KBr pellets. Nuclear magnetic resonance (NMR) spectra
were recorded on a Bruker ARX 400 (400 MHz) spectrometer
F igu r e 4. Summary of results of H9 cell infections by HIV-1
IIIB in the presence of compound 4c: (black) day 7, (light gray)
day 11, and (dark gray) day 14.
using samples in CDCl
3
solution with TMS internal standard
Compound 4c appeared to be the most effective at
inhibiting virus production with almost complete sup-
pression at 100 µM (Figure 4) as well as being the only
compound to significantly delay the onset of syncytial
giant cells in culture (Figure 5). Nucleosides 4e,c were
then compared to the standard anti-HIV agents AZT
and ddI for inhibition of HIV-1 replication (Figure 6).
As shown in Figure 6, the inhibitory effect of compound
or D O with TSP as internal standard. Mass spectra (MS)
2
were recorded at 70 eV on a Kratos profile mass spectrometer.
High-resolution mass spectra (HRMS) were obtained at the
McMaster Regional Centre for Mass Spectrometry using a VG
ZAB-E instrument. Optical rotations were determined using
a Perkin Elmer 24 polarimeter at a wavelength of 589 mm
with a 1.0 dm cell containing a total volume of 1 mL.
Photolysis were performed using a Hanovia 450 W medium
pressure mercury arc lamp in a water-cooled quartz immersion
well. Pyrex test tubes containing the samples were strapped
around this well, and the assembly was immersed in an ice-
water bath. The samples were degassed for 30 min prior to
irradiation. All solvents used were dried and distilled. Mi-
croanalyses were carried out by Guelph Chemical Laboratories
Ltd. The X-ray crystal structure determination was performed
on a Siemens R3m/v diffractometer using a total of 1706
reflections. Preparative HPLC was performed using a Waters
RCM 25 × 10 cartridge at the University of Toronto. Ketone
4
1
c is very similar to that observed for ddI and AZT at
0 µM.
In summary, adenosine analogue 4c is not toxic to
H9 cells at the concentration up to 100 µM and
significantly inhibits HIV-1 replication in an acute
infection system. Further studies of this compound
would include examining its antiviral effect in human
peripheral blood lymphocytes following infection with

Notes
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 26 5279
6
1
was prepared by a modified procedure as described in the
9-[2′,3′-Did eoxy-3′-C-(h ydr oxym eth yl)-r- a n d -â-D-er yth -
r o-p en tofu r a n osyl]p u r in e (5a a n d 4a ). Eluting solvent for
preparative TLC was ethyl acetate:methanol (8:2). Total
yield: 88% total (1:1) each of 4a and 5a .
1
3
literature.
The â-alkoxypurines were prepared by known
4
methods from purin-6-yltrimethylammonium chloride (Ald-
rich Chemical Co.). Anomeric mixtures of protected nucleo-
sides 2a ,b were prepared according to a procedure reported
2
2
r-An om er 5a : mp 161-163 °C; [R]
D
+ 17° (c 0.5, H O);
2
6
1
by us recently.
UV (H O) λ
264 nm (ꢀ ) 8209); H NMR (D O) δ 9.08 (s,
2
max
2
1
-N-(6-Meth oxyp u r in e-9-yl)-5-O-ben zoyl-3-C-[(ben zoyl-
oxy)m eth yl]-2,3-d id eoxy-r- a n d -â-D-er yth r o-fu r a n osid e
2d ). A solution consisting of ketone 1 (136 mg, 0.4 mmol)
and 6-methoxypurine (180 mg, 1.2 mmol) in 150 mL of
acetonitrile was irradiated for 36 h. Evaporation of the solvent
followed by preparative thin-layer chromatography (CH
OH, 95:5) gave 14 mg (12%) of 3 and 81 mg (41%) of the
title compound as a pale yellow solid: IR 1715, 1652 cm ; H
NMR δ 8.50, 8.48 (2 s, 1H), 8.15 (s, 1H), 8.07-8.01 and 7.65-
.53 and 7.50-7.38 (m, 10H), 6.40-6.37 (m, 1H), 4.84-4.40
m, 5H), 4.18 and 4.17 (2 s, 3H), 3.08-2.88 (m, 2H), 2.66-
.56 (m, 1H).
-N-[6-(H exyloxy)p u r in -9-yl]-5-O-b en zoyl-3-C-[(b en -
zoyloxy)m eth yl]-2,3-d id eoxy-r- a n d -â-er yth r o-fu r a n o-
sid e (2e). A solution consisting of ketone 1 (68 mg, 0.2 mmol)
and 6-(hexyloxy)purine (132 mg, 0.6 mmol) in 160 mL of
acetonitrile was irradiated for 36 h. Evaporation of the solvent
1H), 8.89 (s, 1H), 8.66 (s, 1H), 6.48 (m, 1H), 4.34-4.30 (m, 1H),
3.84-3.80 (d × d, J ) 2.5, 13.0 Hz, 1H), 3.72-3.62 (d × d plus
overlapping d, J ) 5.7, 5.9, 13.0 Hz, 3H), 2.84-2.74 (m, 1H),
(
2.58-2.52 (m, 1H), 2.52-2.47 (m, 1H); 13C NMR (D O, TSP) δ
2
157.2, 154.7, 150.8, 142.3, 123.5, 87.9, 86.2, 65.3, 64.5, 44.4,
2
Cl
2
:
37.4; MS (FAB) m/ z 251 (M + 1).
â-An om er 4a : mp 167-169 °C; [R]
6
CH
3
22
D
3
+3.2° (c 0.25, CH -
-
1
1
1
OH); UV (H
2
O) λmax 264 nm (ꢀ ) 7896); H NMR (D
2
O) δ 9.08
(
1
s, 1H), 8.89 (s, 1H), 8.68 (s, 1H), 6.46 (m, 1H), 4.11-4.08 (m,
7
(
2
H), 3.82-3.78 (d × d, J ) 2.8, 12.7 Hz, 1H), 3.71 (d, J ) 5.7
Hz, 2H), 3.66-3.62 (d × d, J ) 5.0, 12.7 Hz, 1H), 2.77-2.66
13
(
m, 2H), 2.53-2.46 (m, 1H); C NMR (D O, TSP) δ 157.8,
2
1
1
54.8, 150.9, 141.6, 122.4, 87.7, 87.4, 65.0, 43.6, 37.6; MS (FAB)
m/ z 252 (M + 2).
-Ch lor o-9-[2′,3′-d id eoxy-3′-C-(h yd r oxym eth yl)-r- a n d
â-D-er yth r o-p en tofu r a n osyl]p u r in e (5b a n d 4b). Eluting
solvent for preparative TLC was ethyl acetate:methanol (85:
5). Total yield: 19% (∼1:1 mixture of anomers) in addition
to 41% of the 6-methoxy nucleosides 4d and 5d .
6
-
followed by preparative TLC (CH
mg (15%) of 3 and 34 mg (30%) of the title compound.
Further separation by preparative TLC (CH Cl :hexane, 3:2)
2 2
Cl :ethyl acetate, 95:5) gave
1
9
2
2
22
r-An om er : mp 199-201 °C; [R]
OH); UV (H O) λmax 260 nm (ꢀ ) 14 074); H NMR (D
.27 (s, 1H), 8.13 (s, 1H), 6.28 (m, 1H), 4.26-4.20 (m, 1H),
.81-3.77 (d × d, J ) 2.7, 12.6 Hz, 1H), 3.68-3.62 (d × d plus
D
3
+35.2° (c 0.165, CH -
afforded 14 mg (13%) of the â-anomer and 14 mg of the
1
2
2
O) δ
R-anomer.
8
3
r-An om er : IR 1714, 1645 cm-1; 1H NMR δ 8.94 (s, 1H), 8.15
s, 1H), 8.07-7.95 and 7.62-7.40 (m, 10H), 6.41-6.39 (m, 1H),
.75-4.49 (m, 5H), 4.48-4.44 (t, 2H), 3.26-3.22 (m, 1H), 3.05-
.98 (m, 1H), 2.62-2.56 (m, 1H), 1.93-1.87 (m, 2H), 1.55-
.50 and 1.38-1.35 (m, 6H), 0.92 (t, 3H); MS (FAB) m/ z 559
(
overlapping d, J ) 5.7, 5.9, 12.6 Hz, 3H), 2.78-2.70 (m, 1H),
4
2
1
13
2
1
3
.55-2.48 (m, 1H), 2.42-2.35 (m, 1H); C NMR (D O, TSP) δ
2
58.4, 155.5, 151.2, 142.7, 121.7, 87.5, 85.9, 65.4, 64.6, 44.3,
7.4; MS (FAB) m/ z 285 (M + 1).
(M + 1).
â-An om er : mp 198-200 °C; [R]22
-12.7° (c 0.275, CH
OH);
O) δ 8.25 (s,
H), 8.10 (s, 1H), 6.26 (m, 1H), 4.07-4.03 (m, 1H), 3.81-3.76
â-An om er : IR 1715, 1652 cm-1; 1H NMR δ 8.47 (s, 1H), 8.08
s, 1H), 8.07-8.03 and 7.61-7.44 (m, 10H), 6.39-6.37 (m, 1H),
.86-4.56 (m, 5H), 4.54-4.50 (t, 2H), 3.01-2.81 (m, 3H), 1.93-
.87 (m, 2H), 1.53-1.35 (m, 6H), 0.93-0.90 (t, 3H); MS (FAB)
D
3
1
UV (H
1
2
O) λmax 260 nm (ꢀ ) 13 468); H NMR (D
2
(
4
1
(
d × d, J ) 2.7, 12.7 Hz, 1H), 3.69-3.68 (d, J ) 5.6 Hz, 2H),
3
2
1
.63-3.59 (d × d, J ) 5.0, 12.7 Hz, 1H), 2.64-2.57 (m, 2H),
.46-2.40 (m, 1H); 13C NMR (D
O, TSP) δ 158.3, 155.3, 151.2,
42.8, 121.9, 87.4, 87.1, 65.5, 64.9, 37.6; MS (FAB) m/ z 284
m/ z 559 (M + 1)
2
9
-[2′,3′-Dideoxy-3′-C-(h ydr oxym eth yl)-r- a n d -â-D-er yth -
r o-p en tofu r a n osyl)a d en in e (5 a n d 4). The identical proce-
dure was used as described by Sammuelsson. A solution
consisting of 220 mg (0.46 mmol) of 2b was treated with 15
mL of saturated methanolic ammonia (precooled in liquid N )
2
at 100 °C in a sealed thick wall tube for 24 h. The solvent
was removed, and the residue was dissolved in water and
(M), 286 (M + 2), 155 (6-chloropurine +1).
8
6
6-Meth oxy-9-[2′,3′-dideoxy-3′-C-(h ydr oxym eth yl)-r- an d
-
â-D-er yth r o-p en tofu r a n osyl]p u r in e (5d a n d 4d ). Eluting
solvent for preparative TLC was ethyl acetate:methanol (95:
5
). Total yield 72% (1:1 mixture of anomers).
r-An om er : mp 121-123.5 °C; [R]22
+3.5° (c 0.73, CH
OH);
O) δ 8.37 (s,
D
3
washed with CH
concentrated to a small volume, and the residue was purified
by reverse phase column chromatography (H O:CH OH, 9:1)
followed by preparative HPLC (H O:CH OH:CH CN, 58:37:
2
Cl
2
(3 × 5 mL). The aqueous layer was
1
UV (H
1
3
2
O) λmax 252 nm (ꢀ ) 12 380); H NMR (D
2
H), 8.34 (s, 1H), 6.33 (m, 1H), 4.25-4.18 (m, 1H), 4.05 (s, 3H),
.76-3.72 (d × d, J ) 2.7, 12.6 Hz, 1H), 3.63-3.58 (d × d plus
2
3
2
3
3
overlapping d, J ) 5.7, 5.9, 12.6 Hz, 3H), 2.77-2.68 (m, 1H),
2
160.1, 155.0, 153.4, 144.5, 124.0, 87.9, 86.1, 65.3, 64.6, 57.9,
5
4
). The R-anomer 5c was eluted first followed by the â-anomer
c. Yields: 44 mg (23%) of 5c and 61 mg (33%) of 4c.
.52-2.43 (m, 1H), 2.42-2.33 (m, 1H); 13C NMR (D
O, TSP) δ
2
8
22
1
r-An om er 5c: mp 166-168 °C (no lit. mp reported ); [R]
D
44.3, 37.4; MS (FAB) m/ z 281.
+
41.5° (c 0.135, H
2
O); UV (H
2
O) λmax 260 nm (ꢀ ) 12 050); H
â-An om er : mp 138-140 °C; [R]22
-6.7° (c 0.75, CH
OH);
O) δ 8.36 (s,
H), 8.30 (s, 1H), 6.30 (m, 1H), 4.04 (s, 3H), 4.00-3.97 (m, 1H),
NMR (D
2
O, TSP) δ 8.33 (s, 1H), 8.19 (s, 1H), 6.35 (m, 1H),
D
3
1
UV (H
1
2
O) λmax 252 nm (ꢀ ) 9771); H NMR (D
2
4
3
2
(
6
.31 (m, 1H), 3.89-3.84 (d × d, J ) 2.7, 13.0 Hz, 1H), 3.75-
.71 (d × d and overlapping d, J ) 5.5, 13.0, 5.9 Hz, 3H), 2.88-
.77 (m, 1H), 2.68-2.54 (m, 1H), 2.50-2.40 (m, 1H); 13C NMR
3.76-3.70 (d × d, J ) 2.7 Hz, 1H), 3.63-3.61 (d, J ) 5.6 Hz,
2
2
H), 3.56-3.50 (d × d, J ) 5.0, 12.5 Hz, 1H), 2.64-2.50 (m,
2
D O, TSP) δ 158.2, 155.2, 151.0, 142.3, 121.5, 85.0, 83.9, 65.3,
H), 2.43-2.34 (m, 1H); 13C NMR (D
O, TSP) δ 160.1, 154.9,
4.8, 43.9, 37.5; MS (FAB) m/ z 266 (M + 1).
2
17
153.2, 144.7, 123.9, 87.7, 87.2, 65.4, 64.9, 57.8, 43.3, 37.5; MS
(FAB) m/ z 281 (M + 1), 151 (6-methoxypurine + 1).
6-(Hexyloxy)-9-[2′,3′-d id eoxy-3′-C-(h yd r oxym eth yl)-r-
an d -â-D-er yth r o-pen tofu r an osyl]pu r in e (5e an d 4e). Elut-
ing solvent for preparative TLC was ethyl acetate:methanol
(95:5). Total yield: 84% (1:1 mixture of anomers).
â-An om er 4c: mp 192.5-194 °C (lit. mp 192-193 °C);
2
2
[
R]
D
-23° (c 0.135, H
2 2
O); UV (H O) λmax 260 nm (ꢀ ) 10 650);
1
H NMR (D O) δ 8.32 (s, 1H), 8.14 (s, 1H), 6.32 (m, 1H), 4.13
2
(
m, 1H), 3.90-3.87 (d × d, J ) 2.5, 12.6 Hz, 1H), 3.77 (d, J )
5
2
1
.3 Hz, 2H), 3.76-3.71 (d × d, J ) 4.7, 12.6 Hz, 1H), 2.72-
13
2
.61 (m, 2H), 2.58-2.45 (m, 1H); C NMR (D
O, TSP) δ 158.1,
r-An om er : mp 98-100 °C; [R]22
+32.5° (c 0.6, CH
OH);
O) δ 8.42 (s,
55.2, 150.9, 142.6, 121.4, 85.3, 85.0, 65.4, 64.9, 43.4, 37.6; MS
D
3
1
(FAB) m/ z 266 (M + 1).
UV (H
2
O) λmax 252 nm (ꢀ ) 12 718); H NMR (D
2
Deben zoyla tion of P r otected Nu cleosid es 2a -e Gen -
1H), 8.40 (s, 1H), 6.38 (m, 1H), 4.53-4.50 (t, 2H), 4.30-4.25
(m, 1H), 3.82-3.78 (d × d, J ) 2.5, 12.6 Hz, 1H), 3.68-3.63 (d
× d plus overlapping d, J ) 5.3, 5.8, 12.6 Hz, 3H), 2.82-2.75
(m, 1H), 2.59-2.51 (m, 1H), 2.48-2.41 (m, 1H), 1.82-1.78 (m,
2H), 1.45-1.41 (m, 2H), 1.32-1.24 (m, 4H), 0.82-0.78 (t, 3H);
er a l P r oced u r e. Approximately 0.15 mmol of protected
nucleoside 2 was treated with 10 mL of saturated methanolic
ammonia for 24 h at room temperature. The solvent was
evaporated and the residue dissolved in water and washed
1
3
with CH (3 × 3 mL). After evaporation of water in vacuo
below 40 °C, the residue was purified by preparative TLC
eluting solvent and yields specified below).
2
Cl
2
2
C NMR (D O, TSP) δ 158.2, 155.2, 151.0, 142.3, 121.5, 87.8,
86.3, 71.1, 65.3, 64.7, 44.2, 37.5, 33.7, 33.6, 31.0, 27.5, 16.0;
(
MS (FAB) m/ z 352 (M + 2), 222.

5
280 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 26
Notes
â-An om er : mp 102-105 °C; [R]2
2
-6.0° (c 0.4, CH
O) δ 8.42 (s,
H), 8.40 (s, 1H), 6.37 (m, 1H), 4.53-4.50 (t, 2H), 4.09-4.04
OH);
D
3
Refer en ces
1
UV (H
1
(
(
2
1
2
O) λmax 252 nm (ꢀ ) 9832); H NMR (D
2
(
1) (a)York University. (b) University of Toronto. (c) Present ad-
dress: Department of Medicinal Chemistry, School of Pharmacy,
m, 1H), 3.82-3.77 (d × d, J ) 2.7, 12.7 Hz, 1H), 3.71-3.69
3
25 Guo He Rd, Shanghai 200433, P.R.C.
d, J ) 5.6 Hz, 1H), 3.64-3.60 (d × d, J ) 5.0, 12.7 Hz, 1H),
(2) Robins, R. K.; Revankar, G. R. In Antiviral Drug Development;
DeClercq, E., Walker, R. T., Eds.; Plenum Press: New York,
1988, pp 11-23.
.71-2.61 (m, 2H), 2.50-2.42 (m, 1H), 1.85-1.79 (m, 2H),
.45-1.39 (m, 2H), 1.30-1.22 (m, 4H), 0.81-0.78 (t, 3H); 13
O, TSP) δ 160.5, 154.7, 153.9, 144.2, 124.3, 87.7, 87.2,
1.3, 65.4, 64.9, 43.4, 37.6, 33.6, 33.5, 30.9, 27.6, 16.1; MS
C
(
3) Pontikis, R.; Wolf, J .; Monneret, C.; Florent, J .-C. Tetrahedron
Lett. 1995, 36, 3523-3526.
NMR (D
2
7
(
(
4) Burns, C. L.; St. Clair, M. H.; Frick, L. W.; Spector, T.; Averett,
D. R.; English, M. L.; Holmes, T. J .; Krenitsky, T. A.; Koszalka,
G. W. J . Med. Chem. 1993, 36, 378-384.
FAB) m/ z 351 (M + 1), 221 (6-(hexyloxy)purine + 1).
In h ibition Stu d ies. The CD4 + T-lymphoblastoid H9 cell
line (NCI, Bethesda, MD) was grown in RPMI 1640 medium,
supplemented with 20% heat-inactivated fetal bovine serum
(5) Lee-Ruff, E.; J iang, J . L.; Wan, W.-Q. Tetrahedron Lett. 1993,
34, 261-264.
(
6) Lee-Ruff, E.; Wan, W.-Q.; J iang, J .-L. J . Org. Chem. 1994, 59,
(
Sigma Chemicals, St. Louis, MO), penicillin (250 µg/mL),
2
114-2118.
(7) Lee-Ruff, E.; Xi, F.; Qie, J .-H. J . Org. Chem. 1996, 61, 1547-
550.
streptomycin (250 µg/mL), L-glutamine (2 mM), and Hepes
6
buffer (10 mM). Cells (2 × 10 /2 mL) were cultured in 12-
1
well tissue culture dishes (Costar, Cambridge, MA). The
adenosine analogues were dissolved in PBS. Cells were
cultured for up to 14 days in the presence of various analogues
tested at 1, 50, and 100 µM and examined for the presence of
cytotoxicity by trypan blue exclusion (0.4%; Gibco, Grand
Island, NY). Acute infections were performed by adding 500
(
8) Svansson, L.; Kvarnstr o¨ m, I.; Classon, B.; Sammuelsson, B. J .
Org. Chem. 1991, 56, 2993-2997.
(9) Seki, J . I.; Shimada, K.; Takahashi, K.; Takita, T.; Takeushi,
T.; Hoshino, H. Antimicrob. Agents Chemother. 1989, 33, 773-
7
75.
(
10) Nishiyama, Y.; Yamamoto, N.; Takahashi, K.; Shimada, N.
Antimicrob. Agents Chemother. 1988, 32, 1053-1056.
11) Fischl, M. A.; Richman, D. D.; Grieco, M. H.; Gottlieb, M. S.;
Volberding, P. A.; Laskin, O. L.; Leedom, J . M.; Groopman, J .
E.; Mildvan, D.; Schooley, R. T. N. Engl. J . Med. 1987, 317, 185-
191.
6
TCID50 HIV IIIB/10 cells to cultures in the presence or
absence of nucleoside, with no subsequent wash step. Cells
were examined for the presence of syncytial giant cells every
(
2
days. Medium was changed at least twice per week.
Supernatants were collected on days 7, 11, and 14 and
examined for p24 antigen using the Abbot ELISA kit according
to the manufacturer’s guidelines.
(12) Nye, K. E.; Knox, K. A.; Pinching, A. J . AIDS 1991, 5, 413-417.
(
(
(
13) Ahmad, S. Tetrahedron Lett. 1991, 32, 6997-7000. (b) Ahmad,
S. Eur. Patent 0458643A2, 1991.
14) Okabe, M.; Sun, R.-C.; Tam, Y.-K.; Todaro, J . L.; Coffen, D. L.
J . Org. Chem. 1988, 53, 4780-4786.
Ack n ow led gm en t. The authors would like to thank
the Natural Sciences and Engineering Research Council
of Canada, the National Health and Research Develop-
ment Program, and the Medical Research Council for
financial support of this work. AZT and ddI were
provided by Dr. Tony Mezzuli (Mount Sinai Hospital,
Toronto).
15) Schneider, K. C.; Benner, S. A. Tetrahedron Lett. 1990, 31, 335-
338.
(
(
16) Sequin, V.; Tamm, C. Helv. Chim. Acta 1972, 55, 1196-1218.
17) Tseng, C. K.-H.; Marquez, G. W. A.; Miline, W. A.; Wysock, R.
J ., J r.; Mitsuya, H.; Shirasaki, T.; Driscoll, J . S. J . Med. Chem.
1991, 34, 343-349.
J M950822K