Synthesis of 2-modified aristeromycins and their analogs as potent inhibitors against Plasmodium falciparum S-adenosyl-l-homocysteine hydrolase

Article abstract of DOI:10.1016/j.bmc.2008.01.046

2-Modified aristeromycin derivatives and their related analogs were synthesized to investigate their inhibitory activity against human and Plasmodium falciparum S-adenosyl-l-homocysteine hydrolase (PfSAHH). 2-Fluoroaristeromycin showed a strong inhibitory activity against PfSAHH selectively and complete resistance to adenosine deaminase.

Full text of DOI:10.1016/j.bmc.2008.01.046

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry 16 (2008) 3809–3815
Synthesis of 2-modified aristeromycins and their analogs
as potent inhibitors against Plasmodium falciparum
S-adenosyl-L-homocysteine hydrolase
Takayuki Ando,a,b Masafumi Iwata,b Fazila Zulfiqar,b Tatsuya Miyamoto,b
Masayuki Nakanishib and Yukio Kitadea,b,c,d,
*
aCenter for Emerging Infectious Diseases, Gifu University, Yanagido 1-1, Gifu 501-1193, Japan
bDepartment of Biomolecular Science, Faculty of Engineering, Gifu University, Yanagido 1-1, Gifu 501-1193, Japan
cCenter for Advanced Drug Research, Gifu University, Yanagido 1-1, Gifu 501-1193, Japan
dUnited Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, Yanagido 1-1, Gifu 501-1193, Japan
Received 30 November 2007; revised 24 January 2008; accepted 25 January 2008
Available online 30 January 2008
Abstract—2-Modified aristeromycin derivatives and their related analogs were synthesized to investigate their inhibitory activity
against human and Plasmodium falciparum S-adenosyl-L-homocysteine hydrolase (PfSAHH). 2-Fluoroaristeromycin showed a
strong inhibitory activity against PfSAHH selectively and complete resistance to adenosine deaminase.
ꢀ 2008 Elsevier Ltd. All rights reserved.
1. Introduction
product possessing inhibitory activity against SAHH.1
However, aristeromycin is deaminated by adenosine
deaminase to yield chemotherapeutically inactive ino-
sine congeners, reducing its inhibitory potency. It has
been recognized that the introduction of a halogen atom
at the 2-position of an adenosine nucleoside enhances its
resistance to the adenosine deaminase.8,9 Actually, 2-flu-
oroneplanocin A (1) exhibited complete resistance to
this enzyme.10
S-Adenosyl-L-homocysteine (SAH) is formed by the
donation of the methyl group of S-adenosyl-L-methio-
nine (SAM) to a methyl acceptor, and is physiologically
hydrolyzed to adenosine and homocysteine by SAH
hydrolase (SAHH). Therefore, the inhibition of SAHH
results in cellular accumulation of SAH, which is a po-
tent feedback inhibition of SAM-dependent biological
methylation such as that at the cap structure at 50-end
of eucaryotic m-RNA.1–4 The eucaryotic m-RNA must
possess a methylated 50-cap structure for stability
against phosphatases and ribonuclease, proper binding
to ribosomes and the promotion of splicing. Therefore,
an uncapped m-RNA is much less likely to be translated
into its respective protein.1–6 Hence, SAHH has emerged
as a target enzyme for the molecular design of the
above-mentioned chemotherapeutic agents.
Furthermore, we have reported that 2-fluoronoraristero-
mycin (2) selectively inhibits Plasmodium falciparum
SAHH (PfSAHH) which has additional space near 2-
position of the adenine-ring, in the substrate binding
pocket, in comparison with human SAHH
(HsSAHH).11,12 Mutagenic analysis of the amino acid
residue forming the additional space confirmed that
the inhibition selectivity is due to the difference of only
one amino acid residue.13
Various carbocyclic adenine nucleosides (1–4, 7) as
SAHH inhibitors have been synthesized (Fig. 1).7 It is
known that aristeromycin (4) is a naturally occurring
On the basis of these observations, we designed and syn-
thesized 2-fluoroaristeromycin (5) and its analogs (6, 8,
and 9 in Figure 1; 10 and 11 in Scheme 2; 12 in Scheme
3) to investigate not only the inhibitory activities against
HsSAHH and PfSAHH but also the resistance to aden-
osine deaminase for the development of a potent anti-
malarial agent.
Keywords: 2-Fluoroaristeromycin; S-Adenosyl-L-homocysteine hydro-
lase; Carbocyclic nucleoside; Enzyme inhibitor.
*
Corresponding author. Tel./fax: +81 58 2932640; e-mail addresses:
ykkitade@gifu-u.ac.jp; kitade@biomol.gifu-u.ac.jp
0968-0896/$ - see front matter ꢀ 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.01.046

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T. Ando et al. / Bioorg. Med. Chem. 16 (2008) 3809–3815
Figure 1. Various carbocyclic adenine nucleosides.
2. Chemistry
20,30-deoxy-2-fluoroaristeromycin (10). The 20,30-epoxy
compound 19 was obtained by the oxidation of 14 with
A synthetic route for the 2-modified aristeromycins and
their 20,30 epimers is shown in Scheme 1. The cyclopen-
tenyl purine derivatives 14 and 15 were prepared by the
coupling reaction of 1314 with 2-fluoroadenine and 2-
amino-6-chloropurine, respectively, in the presence of
a palladium catalyst.15 The treatment of 14 and 15 with
methanolic ammonia yielded the de-acylated and/or 6-
amidated compounds 16 and 17, respectively. Osmium
oxidation16 of these compounds with N-methylmorpho-
rine N-oxide (NMO) yielded 2-fluoroaristeromycin, 2-
aminoaristeromycin, and their 20,30 epimers (5, 6, 8,
and 9).
m-chloroperbenzoic acid (m-CPBA),17 and the subse-
18,19
quent reduction using LiAlH4
30-deoxy compound 11 (Scheme 2).
afforded the 20-epi-
Compound 16 was oxidized by m-CPBA to afford 20,30-
epoxy-2-fluoroaristeromycin (12, Scheme 3). Although it
was possible to obtain 12 from 19 by de-acetylation,
ring-opening compounds were obtained as inseparable
by-products along with the desired epoxy compound.
The structures of these compounds were supported by
spectrum data (1H NMR, 13C NMR, HREIMS). The
stereochemical assignments were mainly based on the
differential NOE correlation of the corresponding
protons.
The catalytic hydrogenation of 14 afforded the reduced
compound 18; subsequent de-acetylation afforded
Scheme 1. Reagents and conditions: (a) 2-fluoroadenine or 2-amino-6-chloropurine, NaH, DMSO, then (Ph3P)4Pd, Ph3P, THF, 55 ꢁC, 73% (for 14)
or 46% (for 15); (b) NH3/MeOH, 0 ꢁC, 99% (for 16) or 120 ꢁC, 71% (for 17); (c) OsO4, NMO, THF–H2O, rt, 45%, 52%, 35% and 40% (for 5, 6, 8 and
9).
Scheme 2. Reagents and conditions: (a) H2, Pd–C, MeOH, rt, 55%; (b) NH3/MeOH, 0 ꢁC, 93%; (c) m-CPBA, CH2Cl2, rt, 56%; (d) LiAlH4, THF, rt,
20%.

T. Ando et al. / Bioorg. Med. Chem. 16 (2008) 3809–3815
3811
4. Discussion
According to the SAHH assay of compounds 5 and 6, it
is clear that the introduction of small molecular, such as
a fluorine atom or a amino residue at the 2-position of
adenine ring, increased the selective indexes, which was
consistent with our previous work.11 The inhibitory
activity of 5 against PfSAHH was better than those of
2-fluoronoraristeromycin (2) because the sugar moiety
structure of these aristeromycin analogs are more simi-
lar to that of adenosine. Furthermore, the deoxy- and
epi-analogs did not show any inhibitory activity. This re-
sult indicates that 20 and/or 30-hydroxyl groups are
essential for the inhibitions against both the SAHHs-
HsSAHH and PfSAHH. The detailed modification of
the sugar moiety is under the investigation.
Scheme 3. Reagents and condition: (a) m-CPBA, CH2Cl2, rt, 77%.
3. Biological assay
3.1. Inhibitory activity against SAHH
The inhibitory activities of the title compounds against
HsSAHH and PfSAHH are summarized as IC50 values
in Table 1. The 2-fluorinated compound 5 and 2-amino
compound 6 showed strong inhibitory activities against
PfSAHH with IC50 value of 1.98 and 4.51 and superior
selective index 24 and 20, respectively; these values were
better than those of the typical compounds we had syn-
thesized. The other synthetic compound in this report
did not show any inhibitory activity up to 500 lM.
On the other hand, 2-fluoroaristeromycin (5) showed the
complete resistance to adenosine deaminase in compar-
ison with that of aristeromycin. This result confirmed
the previous report,10 which stated the introduction of
a halogen atom was effective in designing inhibitors
showing the resistance to adenosine deaminase.
3.2. Susceptibility to adenosine deaminase
In this paper, we have demonstrated the synthesis of 2-
modified aristeromycin analogs and their inhibitory
activity against HsSAHH and PfSAHH. Among the re-
ported compounds, 5 was the best selective inhibitor
against PfSAHH, showing resistance to adenosine
deaminase. These results will significantly contribute to
the design of potent inhibitors as antimalarial agents.
The susceptibility of the compounds to adenosine deam-
inase was investigated under the previously reported con-
dition21 (Table 2). Compound 5 was completely resistant
to adenosine deaminase unlike aristeromycin, which was
rapidly deaminated within 15 min under the reaction
conditions. 2-Aminoaristeromycin was slightly deami-
nated with a 5% conversion ratio.
5. Experimental
Table 1. Inhibitory activities of carbocyclic nucleosides against human
and P. falciparum SAH hydrolases
5.1. General remarks
Compound
HsSAHH
IC50 (lM)
63a
1.1a
PfSAHH
IC50 (lM)
13a
Selective
indexd
1H and 13C NMR spectra were recorded at 400 MHz on
a JEOL AL-400 (operated at 400 and 100 MHz, respec-
tively) by using CDCl3 with TMS as the internal stan-
dard, DMSO-d6, and CD3OD. The spin multiplicities
are indicated by the following symbols s (singlet), d
(doublet), dd (doublet of doublet), t (triplet), ddd (dou-
blet of doublet of doublet), q (quartet), m (multiplet),
and br (broad). Coupling constants (J) are expressed
in Hertz. The differential nuclear overhauser effect (DIF-
NOE) data were described; the irradiated proton (the
correlated proton; % of enhancement value). Mass spec-
tra (HREIMS) were recorded at 70 eV on Shimadzu QP
1000 A spectrometer. Reactions were monitored by thin-
layer chromatography using E. MERCK silica gel 60
F254 glass plate. Silica gel column chromatography was
carried out on Wako gel C-300. HPLC was performed
using the Shimadzu HPLC 10A VP-series attached to
an YMC ODS-AM column (250 mm · 20 mm) for sep-
aration or an YMC ODS-M80 column (150 mm ·
8 mm) for analysis.
2
3
4.8
0.35
0.085
24
3.1a
57.0
1.98
4.51
2.1b
NDc
NDc
NDc
NDc
NDc
4
4.85
47.2
90.7
15.7b
NDc
NDc
NDc
NDc
NDc
5
6
20
7
7.5b
—
8
9
—
—
10
11
12
—
—
a Ref. 11.
b Ref. 7c.
c No inhibitory activity showed at 500 lM.
d Selective index: mean of IC50value for HsSAH/mean of IC50 value for
PfSAHH.
Table 2. Effect of calf intestinal adenosine deaminase on 2-modified
aristeromycins
Compound
15 min
5.1.1. 9-[cis-(10R,40S)-40-Acetoxymethyl-20-cyclopenten-
10-yl]-9-H-2-fluoroadenine (14). A mixture of 2-fluoroad-
enine (385 mg, 2.5 mmol) and 60% NaH (98 mg,
2.46 mmol) in DMSO (5 mL) was stirred at rt for
0.5 h. The reaction mixture was added to a solution of
4
5
6
22.4a
99.5a
95.2a
a % of remaining compound after incubation.

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T. Ando et al. / Bioorg. Med. Chem. 16 (2008) 3809–3815
tetrakis(triphenylphosphine)palladium(0)
(377 mg,
5.1.3. 9-[cis-(10R,40S)-40-Hydroxymethyl-20-cyclopenten-
10-yl]-9-H-2-fluoroadenine (16). Compound 14 (684 mg,
2.35 mmol) was treated with methanolic ammonia
(50 mL, 27% w/w) at 0 ꢁC for 36 h. The reaction mixture
was evaporated under reduced pressure. Column chro-
matography (CHCl3/MeOH, 15:1) of the residue on sil-
0.33 mmol), triphenylphosphine (86 mg, 0.33 mmol)
and 13 (647 mg, 3.26 mmol) in THF (5 mL) under the
dark condition and then stirred at 55 ꢁC for 24 h. The
reaction mixture was evaporated under reduced pres-
sure. Chloroform was added to the residue and the
insoluble matter was removed by filtration. The filtrate
was washed with brine, dried (Na2SO4) and concen-
trated. Column chromatography (EtOAc/hexane, 3:1)
of the residue on silica gel gave 14 (537 mg, 73%) as
1
ica gel gave 16 (585 mg, 99%) as white solid. H NMR
(DMSO-d6, 400 MHz) d 8.02 (1H, s, H-8), 7.75 (2H,
br s, NH2), 6.13 (1H, m, H-30), 5.89 (1H, m, H-20),
5.46 (1H, m, H-10), 4.71 (1H, t, J6 ;OH ¼ 5:4, OH-60),
0
1
white solid. H NMR (CDCl3, 400 MHz) d 7.80 (1H,
3.45 (2H, m, H-60), 2.87 (1H, m, H-40), 2.65 (1H, dt,
s, H-8), 6.18 (1H, m, H-30), 5.95 (1H, m, H-20), 5.78
(2H, br s, NH2), 5.65 (1H, m, H-10), 4.15 (2H, m, H-
60), 3.18 (1H, m, H-40), 2.91 (1H, m, H-50b), 2.07 (3H,
s, CH3CO), 1.67 (1H, m, H-50a); DIFNOE: H-10 (H-20:
3.9%, H-50a: 8.8%), H-40 (H-30: 1.7%, H-50a: 5.5%, H-60:
2.6%); 13C NMR (CDCl3, 100 MHz) 170.92, 159.12 (d,
J = 210.7), 157.02 (d, J = 20.6), 151.24 (d, J = 18.9),
138.80 (d, J = 2.5), 138.07, 129.83, 118.01 (d, J = 4.1),
66.29, 59.43, 44.45, 35.17, 20.88; HREIMS calcd for
C13H14FN5O2 291.1132. Found: 291.1122.
Jgem = 13.8, H-50 Þ, 1.62 (1H, dt, H-50 ); 13C NMR
a
(DMSO-d6, 100 MHz) 158.51 (d, J = 204.1), 157.55 (d,
b
J = 21.4),
150.40
(d,
J = 20.6),
139.24
(d,
J = 2.5),138.53, 129.36, 117.30 (d, J = 4.1), 63.87,
59.24, 47.75, 34.05; HREIMS calcd for C11H12N5OF:
249.1026. Found: 249.1030; Anal. Calcd for
C11H12N5OFÆ1/10H2O: C, 52.62; H, 4.90; N, 27.90.
Found: C, 52.52; H, 4.81; N, 27.85.
5.1.4. 9-[cis-(10R,40S)-40-Hydroxymethyl-20-cyclopenten-
10-yl]-9-H-2-aminoadenine (17). Compound 15 (400 mg,
1.30 mmol) was treated with methanolic ammonia
(50 mL, 27% w/w) in a sealed tube at 120 ꢁC for 48 h.
The reaction mixture was evaporated under reduced
pressure. Column chromatography (CHCl3/MeOH,
15:1) of the residue on silica gel gave 17 (316 mg, 99%)
1
as white solid. H NMR (DMSO-d6, 100 MHz) d 7.60
(1H, s, H-8), 6.62, 5.73 (4H, 2br s, NH2-2, NH2-6),
6.09 (1H, m, H-30), 5.86 (1H, m, H-20), 5.36 (1H, m,
H-10), 4.73 (1H, t, OH-60), 3.43 (2H, m, H-60), 2.85
(1H, m, H-40), 2.57 (1H, m, Jgem = 13.8, H-50a), 1.57
(1H, m, H-50b); DIFNOE: H-10 (H-20: 5.5%, H-50a:
6.8%, H-8: 2.4%), H-40 (H-30: 5.5%, H-60: 4.5%, H-50a:
3.8%); 13C NMR (DMSO-d6, 100 MHz) 160.32,
156.28, 151.58, 138.13, 135.25, 130.22, 113.50, 64.27,
58.31, 47.86, 34.46; HREIMS calcd for C11H14N6O
246.1229. Found: 246.1224.
5.1.2. 9-[cis-(10R,40S)-40-Acetoxymethyl-20-cyclopenten-
10-yl]-9-H-2- amino-6-chloropurine (15). Compound 15
(671 mg, 2.18 mmol) was prepared from 2-amino-6-
chloropurine (2.2 g, 13.0 mmol) and 13 (1 g, 5.05 mmol)
as described from 14 in 43% yield after silica gel chroma-
tography (EtOAc/hexane, 4:1). 1H NMR (CDCl3,
400 MHz) d 7.80 (1H, s, H-8), 6.13 (1H, m, H-30), 5.88
(1H, m, H-20), 5.56 (1H, m, H-10), 5.08 (2H, br s,
NH2), 4.17 (2H, m, H-60), 3.17 (1H, m, H-40), 2.84
(1H, dt, Jgem = 14.4, H-50a), 2.05 (3H, s, CH3CO), 1.69
(1H, m, Jgem = 14.4, H-50b); DIFNOE: H-10 (H-20:
8.6%, H-50a: 14.8%), H-40 (H-30: 3.3%, H-50a: 5.3%, H-
60: 4.9%); 13C NMR (CDCl3, 100 MHz) 170.99,
158.92, 153.35, 151.21, 140.52, 137.92, 129.79, 125.56,
66.14, 59.46, 44.41, 34.53, 20.89; HREIMS calcd for
C13H14N5O2Cl: 307.0836. Found: 307.0845.
5.1.5. 9-[(10R,20S,30R,40R)-20,30-Dihydroxy-40-hydroxy-
methyl-cyclopentan-10-yl]-9-H-2-fluoroadenine (5) and
9-[(10R,20R,30S,40R)-20,30-dihydroxy-40-hydroxymethyl-
cyclopentan-10-yl]-9-H-2-fluoroadenine (8). To a stirred
solution of compound 16 (250 mg, 1.0 mmol) and N-
methylmorpholine N-oxide (NMO, 353 mg, 3.0 mmol)
in THF (10 mL) and H2O (1.0 mL), 2% OsO4solution
(1.3 mL, 0.10 mmol) was added and stirred at rt for
6 h. The mixture was evaporated under reduced pres-
sure. Column chromatography (stepwise elution
CHCl3/MeOH/H2O, 5:1:0 ! 3:1:0 ! 5:4:1) of the resi-
due on silica gel and preparative PLC (H2O/MeOH,
90:10) gave 5 (128 mg, 45%) and 8 (99 mg, 35%) as white
1
solid. For 5: H NMR (DMSO-d6, 400 MHz) d 8.18
(1H, s, H-8), 7.73 (2H, s, NH2-6), 4.91 (1H, d,
J2 ;OH ¼ 6:80, OH-20), 4.70 (1H, t, J6 ;OH ¼ 5:36, OH-
0
0
0
0
60), 4.65 (1H, d, J3 ;OH ¼ 4:16, OH-30), 4.58 (1H, m,
H-10), 4.25 (1H, m, H-20), 3.80 (1H, m, H-30), 3.43
(2H, m, H-60), 2.20 (1H, m, H-50a), 2.01 (1H, m, H-40),
1.63 (1H, m, H-5b0 ); DIFNOE: H-10 (H-50a: 8.3%), H-30
(H-20: 1.4%, H-5b0 : 0.4%), H-40 (H-30: 3.0%, H-50a:
3.1%); 13C NMR (DMSO-d6, 100 MHz) 158.27 (d,
J = 203.3), 157.51 (d, J = 21.4), 150.95 (d, J = 19.8),
0
0

T. Ando et al. / Bioorg. Med. Chem. 16 (2008) 3809–3815
3813
140.34, 117.56, 74.57, 71.53, 62.96, 59.15, 45.26, 29.15;
HREIMS calcd for C11H14FN5O3 283.1081. Found:
283.1086; Anal. Calcd for C11H14FN5O3Æ1/10H2O: C,
46.35; H, 5.02; N, 24.57. Found: C, 45.95; H, 4.84; N,
24.66. For 8: H NMR (DMSO-d6, 400 MHz) d 8.14
(1H, s, H-8), 7.70 (2H, s, NH2-6), 5.01, 4.93 (2H, d,
156.72, 152.64, 138.60, 113.49, 72.76, 72.39, 61.33,
53.36, 40.08, 33.09; HREIMS calcd for C11H16FN6O3
280.1284. Found: 280.1291.
1
J
2 ;OH, J3 ;OH ¼ 5:6, 3.92, OH-20,30), 4.80 (1H, m, H-10),
0
0
4.48 (1H, t, J6 ;OH ¼ 5:36, OH-60), 4.50 (2H, s, H-20
0
and 30), 3.55 (2H, m, H-60), 2.24 (1H, dt, J1 ;5 a ¼ 8:3,
Jgem = 12.7, H-50a), 2.05 (1H, m, H-40), 1.81 (1H, m,
H-5b0 ); DIFNOE: H-10 (H-20: 9.2%, H-50a: 5.6%), H-40
(H-30: 6.8%, H-5a0 : 3.3%); 13C NMR (DMSO-d6,
100 MHz) d 158.45 (d, J = 204.1), 157.37 (d, J = 21.4),
151.23 (d, J = 20.6), 141.71 (d, J = 2.5), 116.45 (d,
J = 4.1), 71.73 (d, J = 43.6), 60.46, 53.19, 48.58, 41.47,
32.83; HREIMS calcd for C11H14FN5O3 283.1081.
Found: 283.1087; Anal. Calcd for C11H14FN5O3: C,
46.64; H, 4.98; N, 24.72. Found: C, 46.41; H, 4.83; N,
24.80.
0
0
5.1.7. 9-[cis-(10R,30S)-10-Acetoxymethyl-cyclopentan-30-
yl]-9-H-2-fluoroadenine (18). Compound 14 (60 mg,
206 lmol) in MeOH (20 mL) was hydrogenated in the
presence of 5% Pd on carbon (50 mg) for 24 h at rt
and the reaction mixture was filtered through Celite,
washing with MeOH. The filtrate was concentrated,
and column chromatography (EtOAc/hexane, 8:3) of
the residue on silica gel gave 10 (33 mg, 55%) as white
1
solid. H NMR (CDCl3, 400 MHz) d 7.83 (1H, s, H-
8), 6.29 (2H, s, NH2), 4.81 (1H, m, H-10), 4.13 (2H, m,
H-60), 2.48–1.76 (7H, m, H-20, 30, 40, and 50), 2.08 (3H,
s, CH3CO); 13C NMR (CDCl3, 100 MHz) 171.15,
161.29 (d, J = 256.8), 156.96 (d, J = 20.6), 151.57 (d,
J = 19.8), 138.80 (d, J = 3.3), 118.15 (d, J = 4.1), 67.39,
55.62, 36.82, 36.11, 31.49, 26.74, 20.94; HREIMS calcd
for C13H16FN5O2 293.1288. Found: 293.1293.
5.1.8. 9-[cis-(10R,30S)-10-Hydroxymethyl-cyclopentan-30-
yl]-9-H-2-fluoroadenine (10). Compound 10 (102 mg,
406 lmol) was prepared from 18 (128 mg, 436 lmol) as
described from 16 in 93% after silica gel chromatogra-
phy (CHCl3/MeOH, 12:1). 1H NMR (DMSO-d6,
400 MHz) d 8.20 (1H, s, H-8), 7.74 (2H, br s, NH2),
4.68 (1H, m, H-10), 4.61 (1H, t, OH-60), 3.40 (2H, m,
H-60), 2.28–1.60 (7H, m, H-20, 30, 40, and 50); 13C
NMR (DMSO-d6, 100 MHz) 158.38 (d, J = 204.1),
157.55 (d, J = 21.4), 150.65 (d, J = 20.5), 139.65 (d,
J = 3.3), 117.46 (d, J = 4.1), 64.73, 55.19, 48.59, 35.28,
30.92, 26.25; HREIMS calcd for C11H14FN5O
251.1182. Found: 251.1191.
5.1.6. 9-[(10R,20S,30R,40R)-20,30-Dihydroxy-40-hydroxy-
methyl-cyclopentan-10-yl]-9-H-2-aminoadenine (6) and
9-[(10R,20R,30S,40R)-20,30-dihydroxy-40-hydroxymethyl-
cyclopentan-10-yl]-9-H-2-aminoadenine (9). Compound 6
(148 mg, 0.53 mmol) and 9 (114 mg, 407 lmol) were pre-
pared from 17 (268 mg, 1.1 mmol) as described from 16
in 48% and 37% yield, respectively, after silica gel chro-
matography (stepwise elution CHCl3/MeOH/H2O,
5:1:0 ! 3:1:0 ! 60:25:4) and preparative HPLC (H2O/
1
MeOH, 90:10). For 6: H NMR (DMSO-d6, 400 MHz)
d 7.76 (1H, s, H-8), 6.59, 5.67 (4H, 2br s, 2NH2), 4.94
(1H, d, J2 ;OH ¼ 6:3, OH-20), 4.69 (1H, t, J6 ;OH ¼ 5:12,
OH-60), 4.53–4.48 (2H, m, H-10, OH-30), 4.22 (1H, m,
H-20), 3.79 (1H, m, H-30), 3.45 (2H, m, H-60), 2.018
(1H, m, H-50b), 1.98 (1H, m, H-40), 1.55 (1H, m, H-5a);
DIFNOE: H-30 (H-20: 2.8%, H-40: 6.0%), H-50b (H-20:
0.4%, H-30: 0.2%, H-50a: 3.8%); 13C NMR (DMSO-d6,
100 MHz) d 159.52, 155.73, 151.68, 135.76, 113.20,
74.33, 71.45, 62.66, 57.78, 44.90, 29.24; HREIMS calcd
for C11H16FN6O3 280.1284. Found: 280.1289. For 9: 1H
NMR (DMSO-d6, 400 MHz) d 7.75 (1H, s, H-8), 6.59,
0
0
5.1.9. 9-[(10R,20R,30S,40R)-40-Acetoxymethyl-20,30-epoxy-
cyclopentan-10-yl]-9-H-20-fluoroadenine (19). To a solu-
tion of 14 (100 mg, 343 lmol) in CH2Cl2 (6 mL) was
added 3-chloroperoxybenzoic acid (m-CPBA, 82 mg,
475 lmol), and the mixture was stirred for 18 h at rt.
After completion of the reaction, the mixture was con-
centrated, and extracted with EtOAc. The extract was
washed with sat NaHCO3, and water, dried (Na2SO4),
and concentrated. Column chromatography (EtOAc/
hexane, 8:3) of residue on silica gel gave 19 (59 mg,
0
5.67 (4H, 2br s, 2NH2), 5.02 (1H, d, J3 ;OH ¼ 4:8, OH-
30), 4.91 (1H, d, J2 ;OH ¼ 6:0, OH-20), 4.69 (1H, m, H-
0
10), 4.47 (1H, t, J6 ;OH ¼ 5:0, OH-60), 4.04 (1H, m, H-
0
1
20), 3.99 (1H, m, H-30), 3.63, 3.48 (2H, m, H-60), 2.15
56%) as white crystal. H NMR (CDCl3, 400 MHz) d
(1H, dt, Jgem = 12.4, J1 ;5a ¼ 8:0, H-50a), 2.05 (1H, m,
0
8.08 (1H, s, H-8), 6.03 (2H, br s, NH2-6), 5.03 (1H, t,
H-40), 1.84 (1H, m, J1 ;5 b ¼ 10:4, H-50b); DIFNOE: H-
40 (H-30: 14%, H-50a: 14%); H-30 (H-20: 2.8%, H-50a:
1.8%); 13C NMR (DMSO-d6, 100 MHz) 160.55,
J1 ;5 a ¼ 10; H-10), 4.22 (2H, m, H-60), 3.78 (1H, s, H-
20), 3.68 (1H, s, H-30), 2.54 (1H, m, H-40), 2.34 (1H,
m, H-50a), 2.08 (3H, s, CH3CO), 1.29 (1H, m, H-50b);
0
0
0
0

3814
T. Ando et al. / Bioorg. Med. Chem. 16 (2008) 3809–3815
DIFNOE: H-10 (H-20: 4.1%, H-30: 0.9%, H-50a: 3.0%), H-
20 (H-10: 5.0%, H-30: 6.2%), H-30 (H-10: 0.9%, H-20:
4.6%, H-40: 4.2%); 13C NMR (CDCl3, 100 MHz) d
70.82, 159.23 (d, J = 211.5), 157.06 (d, J = 20.6),
151.43 (d, J = 18.9) 138.93 (d, J = 3.3), 117.35 (d,
J = 4.1), 63.49, 57.22, 56.25, 54.14, 38.36, 28.94, 20.83;
HREIMS calcd for C13H14FN5O3 307.1081. Found:
307.1078.
Jgem = 12.5, H-50a), 1.15 (1H, m, H-50b); DIFNOE: H-10
(H-20: 9.2%, H-5a0 : 7.0%), H-20 (H-10: 5.1%, H-30:
3.4%), H-30 (H-10: 1.4%, H-20: 3.8%), H-40 (H-30:
4.2%, H-50a: 6.3%, H-60: 3.0%); 13C NMR(CDCl3,
100 MHz) 158.68 (d, J = 204.1), 157.60 (d, J = 21.4),
150.78 (d, J = 20.6), 138.86 (d, J = 2.5), 116.94 (d,
J = 4.1), 61.00, 56.94, 56.44, 54.37, 41.26, 27.83; HRE-
IMS calcd for C11H12FN5O2 265.0975. Found:
265.0966.
5.1.10. 9-[(10R,20S,40S)-20-Hydroxy-40-hydroxymethyl-
cyclopentan-10-yl]-9-H-2-fluoroadenine (11). To a solu-
tion of 19 (50 mg, 163 lmol) in THF (5 mL) was added
LiAlH4 (71 mg, 1.9 mmol), and the mixture was stirred
for 5 h at rt. After completion of the reaction, methanol
was added at 0 ꢁC, and the mixture was filtered through
Celite. he filtrate was concentrated and preparative
HPLC (MeOH/H2O, 30:70) of residue gave 11 (10 mg,
5.2. Enzyme assay
In the synthetic direction, the enzyme assay was a mod-
ification of an earlier method.20 The enzyme was incu-
bated with 100 mM adenosine, 5 mM DL-homocysteine
and inhibitors on 0.2 mL of 10 mM potassium phos-
phate, pH 7.2, buffer at 30 ꢁC for 2 min in the standard
assay system. The reaction was started by the addition
of 3 mL of SAH hydrolase (human: 0.43 lg, P. falcipa-
rum: 0.54 lg) and terminated by the addition of 20 lL of
0.67 N HCl. The reaction mixture was kept on ice until
the HPLC analysis. The mixture was analyzed for SAH
by a Shimadzu HPLC system. In the synthetic reaction,
one unit of SAHH was defined as the amount synthesiz-
ing 1 mmol of SAH/min at 30 ꢁC.
1
23%) as white solid. H NMR (CD3OD, 400 MHz) d
8.19 (1H, s, H-8), 4.69 (1H, m, H-10), 4.29 (1H, m, H-
20), 3.63 (2H, m, H-60), 2.35–2.23 (3H, m, H-30a, 40,
50a), 2.14 (1H, m, H-5b0 ), 1.62 (1H, m, Jgem = 12.8,
H-30b); DIFNOE: H-20 (H-10: 7.6%, H-3a0 : 1.8%), H-50b
(H-60; 8.1%), H-60 (H-5b0 : 2.4%, H-30b: 2.3%); 13C
NMR (CDCl3, 100 MHz) d158.49 (d, J = 204.1),
157.49 (d, J = 21.4), 151.20 (d, J = 20.6), 140.73 (d,
J = 2.5), 116.78 (d, J = 4.1), 77.20, 64.35, 61.36, 53.84,
46.77, 30.34; HREIMS calcd for C11H14FN5O2
267.1132. Found: 267.1112.
Acknowledgment
This research was in part supported by Grant-in-Aid for
Scientific Research on Priority Area No. 16017239 (to
Y.K.).
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0
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