Does the anti-hepatitis b virus activity of (+)-5'-noraristeromycin exist in its 4'-epimer and 4'-deoxygenated derivatives?

Article abstract of DOI:10.1021/jm980038a

To begin an exploration of the structural parameters responsible for the activity of (+)-5'noraristeromycin toward hepatitis B virus (HBV), three derivatives varied at the C-4' position have been prepared and evaluated. The syntheses began with a Mitsunobu coupling reaction of an appropriate cyclopentanol with 6-chloropurine. The products of these reactions were synthetically altered by standard ammonolysis and deprotection procedures to give the desired products. Evaluation of the new derivatives indicated that removal of the C-4' hydroxyl of (+)-5'-noraristeromycin increased its potency toward HBV by approximately 10-fold.

Full text of DOI:10.1021/jm980038a

2168
J . Med. Chem. 1998, 41, 2168-2170
Notes
Does th e An ti-Hep a titis B Vir u s Activity of (+)-5′-Nor a r ister om ycin Exist in Its
4′-Ep im er a n d 4′-Deoxygen a ted Der iva tives?
Katherine L. Seley and Stewart W. Schneller*
Department of Chemistry, Auburn University, Auburn, Alabama 36849-5312
Brent Korba
Georgetown University Medical Center, Division of Molecular Virology and Immunology, Rockville, Maryland 20852
Received J anuary 21, 1998
To begin an exploration of the structural parameters responsible for the activity of (+)-5′-
noraristeromycin toward hepatitis B virus (HBV), three derivatives varied at the C-4′ position
have been prepared and evaluated. The syntheses began with a Mitsunobu coupling reaction
of an appropriate cyclopentanol with 6-chloropurine. The products of these reactions were
synthetically altered by standard ammonolysis and deprotection procedures to give the desired
products. Evaluation of the new derivatives indicated that removal of the C-4′ hydroxyl of
(+)-5′-noraristeromycin increased its potency toward HBV by approximately 10-fold.
We recently reported1 significant activity for (+)-5′-
Ch a r t 1. Historical Structures and Target Compounds
noraristeromycin (1) (Chart 1) toward hepatitis B virus
(HBV). In exploring derivatives related to 1 that could
possess more potent activity, the methoxy derivative 2
was pursued,2 which indicated that the 4′-hydroxyl
hydrogen of 1 was necessary for its anti-HBV properties.
We then chose to determine the role of the stereochem-
ical configuration of the C-4′ hydroxyl group on the HBV
properties of 1 as well as the importance of the 4′-
hydroxyl itself on the activity. For these purposes, the
epimer 3, the deoxy 4, and the alkene 5 were chosen as
candidate compounds. The results of this study are
described here.
Ch em istr y
The preparation of target compound 3 (Scheme 1)
began with the Mitsunobu coupling of the chiral acetate
63 with 6-chloropurine. This reaction is known to
proceed with inversion of configuration4 and, conse-
quently, led to 7.5 Glycolization of 7 to 8 was followed
by ammonolysis to provide 3.
Presence of a double bond in 4 (that is, 5) resulted in a
slight decrease in potency, but with an accompanying
Scheme 2 shows the synthetic routes used to realize
decrease in toxicity. While none of the derivatives
4 and 5. This pathway also began with a Mitsunobu
described herein are as effective an anti-HBV agent as
is 3TC (Table 1), the results for 4 and 5 suggest them
coupling using the protected glycol cyclopentenol 96 and
6-chloropurine. Ammonolysis of the product of this
as prototype compounds from which new entities should
reaction (10) followed by deprotection with Dowex 50
be designed and studied.
× 8 acidic resin yielded 5.7 Catalytic hydrogenation of
5 resulted in 4.7
Exp er im en ta l Section
Gen er a l. Melting points were recorded on a Meltemp II
melting point apparatus and are uncorrected. Combustion
Resu lts
analyses were performed by M-H-W Laboratories, Phoenix,
The anti-HBV data for compound 3 compared to 1
AZ. 1H and 13C NMR spectra were recorded on a Bruker AC
(Table 1) verified the importance of the original stere-
250 spectrometer (operated at 250 and 62.5 MHz, respectively),
all referenced to internal tetramethylsilane (TMS) at 0.0 ppm.
The spin multiplicities are indicated by the symbols s (singlet),
d (doublet), t (triplet), q (quartet), m (multiplet), and br (broad).
ochemical configuration of the C-4′ hydroxyl group in
1. However, removal of the C-4′ hydroxyl from 1 (as in
4) showed a 7-10-fold increase in anti-HBV activity.
S0022-2623(98)00038-7 CCC: $15.00 © 1998 American Chemical Society
Published on Web 05/01/1998

Notes
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 12 2169
Sch em e 1a
give 10.0 g of 7 contaminated with triphenylphosphine oxide
as a yellow solid which was used directly in the next step
without further characterization.
To the impure 7 (10.0 g) in THF/H2O (11:1, 240 mL) were
added 4-methylmorpholine N-oxide (10 mL) and OsO4 (150
mg), and the solution stirred at room temperature overnight.
The solvents were removed under reduced pressure, the
residue was coevaporated with toluene (2 × 50 mL) and then
acetone (50 mL), and the residue was purified by column
chromatography eluting with CH2Cl2/MeOH (95:5) to afford
2.75 g of 8 (13% from 6) as a white solid: mp 193-194 °C; 1H
NMR (DMSO-d6) δ 2.0 (s, 3H), 2.15 (m, 1H), 2.90 (dt, 1H), 4.12
(m, 2H), 5.17 (br, 1H), 5.23 (m, 1H), 5.34 (m, 1H), 5.37 (t, 1H),
8.05 (s, 1H), 8.79 (br, 2H); 13C NMR (DMSO-d6) δ 20.9, 33.4,
53.5, 71.5, 75.6, 77.4, 130.7, 147.2, 148.7, 151.2, 152.2, 170.3.
Calcd for C12H13Cl N4O4 (C, H, N).
(1S,2R,3R,4S)-4-(6-Am in o-9H-p u r in -9-yl)cyclop en ta n e-
1,2,3-tr iol ((+)-3). A solution of 8 (2.75 g, 8.76 mmol) in
saturated methanolic ammonia was sealed in a steel vessel
and heated at 100 °C for 2 days. The solvent was removed
under reduced pressure, the residue was triturated with H2O
and filtered, and the solvent was removed to give 0.85 g of 9
a
Reaction conditions: (a) 6-chloropurine, PPh3, DIAD, THF; (b)
OsO4/60% aq 4-methylmorpholine N-oxide in THF; (c) NH3 in
MeOH, 120 °C, 2 days.
(39%) as a pale-gray solid: mp 235-236 °C; [R]23 +15.3 ° (c
D
Sch em e 2a
0.59, DMSO); 1H NMR (DMSO-d6) δ 2.01 (t, 1H), 2.46 (m, 1H),
3.83 (br, 1H), 4.11 (m, 2H), 5.06 (m, 4H), 7.18 (br, 2H), 8.12
(s, 1H), 8.17 (s, 1H); 13C NMR (DMSO-d6) δ 37.0, 52.6, 71.6,
73.9, 78.7, 118.3, 140.3, 149.6, 151.9, 155.8. Calcd for
C
10H13N5O3 (C, H, N).
(1′S,2′R,3′S)-9-[2′,3′-(Isop r op ylid en ed ioxy)cyclop en t-4′-
en -1′-yl]-6-ch lor o-9H-p u r in e (10). To a stirring, chilled (-10
°C) suspension of 6-chloropurine (5.66 g, 36.6 mmol) and Ph3P
(9.63 g, 36.7 mmol) in dry THF (100 mL) was added dropwise
DIAD (7.40 g, 36.6 mmol), and the solution stirred for 10 min,
at which point the ice bath was removed and the reaction
stirred for an additional 15 min. To this was added a solution
of 96 (5.20 g, 36.6 mmol) in dry THF (150 mL), and the reaction
mixture stirred at room temperature for 2 h, followed by
stirring at 55 °C for 2 days. The solvent was removed under
reduced pressure and the residue purified via column chro-
matography, eluting with EtOAc, followed by EtOAc/MeOH
(15:1). Fractions containing product were combined and
evaporated to give 2.35 g (24%) of 10 as a white crystalline
a
Reaction conditions: (a) 6-chloropurine, PPh3, DIAD, THF; (b)
NH3 in MeOH, 120 °C, 2 days; (c) Dowex 50 × 8 acidic resin,
MeOH; (d) PtO2, MeOH, H2, 25 psi.
Ta ble 1. Inhibition of Hepatitis B Virus by
5′-Noraristeromycin Derivativesa
1
solid: mp 128-129 °C; H NMR (CDCl3) δ 1.27 (s, 3H), 1.51
SI(CC50
EC90
/
compd
CC50 (µM)
EC50 (µM)
1.4 ( 0.1
EC90 (µM)
9.6 ( 0.8
)
(s, 3H), 4.74 (d, 1H), 5.55 (d, 1H), 5.72 (s, 1H), 5.99 (dd, 1H),
6.42 (d, 1H), 8.01 (s, 1H), 8.78 (s, 1H); 13C NMR (CDCl3) δ 21.9,
25.7, 27.3, 66.2, 83.7, 84.7, 112.7, 128.6, 132.1, 139.1, 143.2,
151.3, 152.2. Calcd for C13H13Cl N4O3 (C, H, N).
1
2
3
4
446 ( 20
>1000
1883 ( 101 >10
93 ( 7.4
325 ( 17
1884 ( 123
46
>10
>10
>10
0.120 ( 0.016
0.145 ( 0.015
0.070 ( 0.008
0.978 ( 0.077
95
232
9014
(1′S,2′R,3′S)-1′-(6-Am in o-9H-p u r in -9-yl)-2′,3′-d ih yd r oxy-
cyclop en t-4′-en e (5). A solution of 10 (2.3 g, 7.86 mmol) in
saturated methanolic ammonia (150 mL) was sealed in a steel
vessel and heated at 110 °C overnight. The solvent was
evaporated, the residue was dissolved in MeOH (100 mL), and
Dowex 50 × 8 resin beads were added. The mixture was
refluxed for 1 h, and the solvent was removed. The residue
was loaded onto a Dowex resin column, and the product was
eluted with concentrated NH4OH. The fractions containing
product were combined and evaporated under reduced pres-
sure. The resultant residue was purified via column chroma-
tography on silica gel eluting with EtOAc and then EtOAc/
MeOH (9:1). Fractions containing the desired product were
combined and evaporated, followed by recrystallization in
MeOH to afford 1.30 g (85%) of 5 as a white solid: mp 180-
5
1.4 ( 0.2
3TC
0.209 ( 0.018
a
For details, see the Experimental Section.
The optical rotations were obtained on a Perkin-Elmer 241
polarimeter. Reactions were monitored by thin-layer chro-
matography (TLC) using 0.25-mm Whatman Diamond silica
gel 60-F254 precoated plates with visualization by irradiation
with a Mineralight UVGL-25 lamp. Column chromatography
was performed on Whatman silica, 230-400 mesh, 60 Å, and
elution with the indicated solvent system. Yields refer to
chromatographically and spectroscopically (1H and 13C NMR)
homogeneous materials.
(1S,2R,3R,4S)-Acetic Acid 4-(6-Ch lor o-9H-p u r in -9-yl)-
2,3-d ih yd r oxycyclop en tyl Ester (8). A stirring, chilled
(-10 °C) suspension of 6-chloropurine (5.0 g, 32.35 mmol) and
Ph3P (8.49 g, 32.35 mmol) in dry THF (100 mL) was treated
dropwise with diisopropyl azodicarboxylate (DIAD) (6.54 g,
32.35 mmol), and the solution stirred at that temperature for
10 min, after which the ice bath was removed and the reaction
stirred for an additional 15 min. To this was then added a
solution of 63 (5.06 g, 35.6 mmol) in dry THF (50 mL), and the
reaction mixture stirred at room temperature for 2 h, followed
by stirring at 55 °C for 2 days. The solvent was evaporated
under reduced pressure, and the residue was purified via
column chromatography eluting with EtOAc/MeOH (9:1) to
181 °C (lit.7c mp 175-176 °C); [R]23 +166.3° (c 0.15, MeOH)
D
1
(lit.7c [R]23 -170.0° (c 1.0, H2O)); H NMR (DMSO-d6) δ 4.33
D
(q, 1H), 4.56 (br, 1H), 4.95 (d, 1H), 5.10 (d, 1H), 5.39 (d, 1H),
5.99 (dd, 1H), 6.13 (m, 1H), 7.19 (s, 2H), 8.08 (s, 1H), 8.12 (s,
1H); 13C NMR (DMSO-d6) δ 64.6, 72.5, 76.1, 119.1, 132.3, 135.9,
139.7, 149.6, 152.2, 155.9. Calcd for C10H11N5O2 (C, H, N).
(1′S,2′R,3′S)-1′-(6-Am in o-9H-p u r in -9-yl)-2′,3′-d ih yd r oxy-
cyclop en ta n e (4). To a solution of 5 (1.0 g, 4.29 mmol) in
MeOH (50 mL) was added PtO2 (0.20 g), and the mixture was
placed under H2 and shaken overnight at 25 psi. The mixture
was filtered over a Celite pad, the pad rinsed with MeOH, and
the filtrate evaporated under reduced pressure. The residue

2170 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 12
Notes
was then purified via column chromatography eluting with
CH2Cl2/MeOH (10:1) (Note: TLC showed no difference from
starting material, but NMR clearly shows the correct product).
Fractions containing product were combined and evaporated
to afford 0.58 g (58%) of 4 as a white crystalline solid: mp
Refer en ces
(1) Seley, K. L.; Schneller, S. W.; Korba, B. A 5′-Noraristeromycin
Enantiomer with Activity Towards Hepatitis B Virus. Nucleo-
sides Nucleotides 1997, 16, 2095-2099.
(2) Seley, K. L.; Schneller, S. W.; De Clercq, E. A Methylated
Derivative of 5′-Noraristeromycin. J . Org. Chem. 1997, 62,
5645-5646.
214-215 °C (lit.7b mp 218 °C); [R]23 +62.1° (c 0.28, MeOH)
D
1
(lit.7b [R]23 -50.0° (c 0.80, H2O)); H NMR (DMSO-d6) δ 1.63
D
(3) Seley, K. L.; Schneller, S. W.; Rattendi, D.; Bacchi, C. J . (+)-7-
Deaza-5′-noraristeromycin as an Anti-trypanosomal Agent. J .
Med. Chem. 1997, 40, 622-624.
(m, 1H), 1.95-2.25 (m, 3H), 4.01 (br, 1H), 4.41 (m, 1H), 4.68
(m, 2H), 4.96 (d, 1H), 7.16 (s, 2H), 8.11 (s, 1H), 8.18 (s, 1H);
13C NMR (DMSO-d6) δ 25.7, 28.8, 59.5, 70.5, 76.1, 119.4, 140.4,
149.6, 151.9, 155.9. Calcd for C10H13N5O2 (C, H, N).
(4) (a) Mitsunobu, O. The Use of Diethyl Azodicarboxylate and
Triphenylphosphine in Synthesis and Transformation of Natural
Products. Synthesis 1981, 1-28. (b) Hughes, D. L. The Mit-
sunobu Reaction. Org. React. 1992, 42, 335-656.
An ti-HBV. Antiviral analyses for the inhibition of HBV
replication were performed on confluent cultures of the chroni-
cally HBV-producing human hepatoblastoma cell line, 2.2.15,
as previously described.8 Cells were treated with nine con-
secutive daily doses (medium removed daily) of the agents.
Antiviral activity was assessed by dot blot hybridization
analysis for reductions in the levels of extracellular HBV virion
DNA and cytotoxicity by uptake of neutral red dye.8 In these
analyses, reductions in virion production of less than 3-fold
are routinely not statistically significant.8
(5) Siddiqi, S. M.; Chen, X.; Schneller, S. W.; Ikeda, S.; Snoeck, R.;
Andrei, G.; Balzarini, J .; De Clercq, E. An Epimer of 5′-
Noraristeromycin and Its Antiviral Properties. J . Med. Chem.
1994, 37, 1382-1384.
(6) (a) Seley, K. L.; Schneller, S. W.; Rattendi, D.; Lane, S.; Bacchi,
C. J . Synthesis and Anti-trypanosomal Activity of Various 8-Aza-
7-deaza-5′-noraristeromycin Derivatives. J . Med. Chem. 1997,
40, 625-629. (b) Seley, K. L.; Schneller, S. W.; Rattendi, D.;
Lane, S.; Bacchi, C. J . Synthesis and Anti-trypanosomal Activi-
ties of a Series of 7-Deaza-5′-noraristeromycin Derivatives with
Variations in the Cyclopentyl Ring Substituents. Antimicrob.
Agents Chemother. 1997, 41, 1658-1661.
(7) (a) Using different synthetic procedures, the enantiomers of 47b
and 57c have been reported. (b) Wolfe, M. S.; Lee, Y.; Bartlett,
W. J .; Borcherding, D. R.; Borchardt, R. T. 4′-Modified Analogues
of Aristeromycin and Neplanocin A: Synthesis and Inhibitory
Activity toward S-Adenosyl-L-homocysteine Hydrolase. J . Med.
Chem. 1992, 35, 1782-1791. (c) Borcherding, D. R.; Scholtz, S.
A.; Borchardt, R. T. Synthesis of Analogues of Neplanocin A:
Utilization of Optically Active Dihydroxycyclopentenones De-
rived from Carbohydrates. J . Org. Chem. 1987, 52, 5457-5461.
(8) Korba, B. E.; Gerin, J . L. Antiviral Res. 1992, 19, 55-70.
Ack n ow led gm en t. This research was supported by
funds from the Department of Health and Human
Services U19-AI31718 (S.W.S.) and also Contract NO1-
AI-45159 (B.K.) between the National Institute of Al-
lergy and Infectious Diseases and Georgetown Univer-
sity, and this is greatly appreciated. We are also grateful
to Dr. J ames Leahy of the University of California at
Berkley for the optical rotation data.
J M980038A