Convenient synthesis of 2′-deoxy-2-fluoroadenosine from 2-fluoroadenine

Article abstract of DOI:10.1081/NCN-120025237

A convenient synthesis of 2′-deoxy-2-fluoroadenosine from commercially available 2-fluoroadenine is described. The coupling reaction of silylated 2-fluoroadenine with phenyl 3,5-bis[O-(t-butyldimethylsilyl)]-2-deoxy-1-thio- D-erythro-pentofuranoside gave

Full text of DOI:10.1081/NCN-120025237

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Convenient Synthesis of 2′-Deoxy-2-fluoroadenosine
from 2-Fluoroadenine
Song Ye a , Martha M. Rezende a , Wei-Ping Deng a & Kenneth L. Kirk a b
a Laboratory of Bioorganic Chemistry , National Institute of Diabetes, and Digestive and
Kidney Diseases, National Institutes of Health, DHHS , Bethesda, Maryland, USA
b Laboratory of Bioorganic Chemistry , National Institute of Diabetes, and Digestive and
Kidney Diseases, National Institutes of Health, DHHS , Bethesda, MD, 20892, USA
Published online: 31 Aug 2006.
To cite this article: Song Ye , Martha M. Rezende , Wei-Ping Deng & Kenneth L. Kirk (2003) Convenient Synthesis of 2′-
Deoxy-2-fluoroadenosine from 2-Fluoroadenine, Nucleosides, Nucleotides and Nucleic Acids, 22:10, 1899-1905, DOI: 10.1081/
NCN-120025237
To link to this article: http://dx.doi.org/10.1081/NCN-120025237
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NUCLEOSIDES, NUCLEOTIDES & NUCLEIC ACIDS
Vol. 22, No. 10, pp. 1899–1905, 2003
Convenient Synthesis of 20-Deoxy-2-fluoroadenosine
from 2-Fluoroadenine
*
Song Ye, Martha M. Rezende, Wei-Ping Deng, and Kenneth L. Kirk
Laboratory of Bioorganic Chemistry, National Institute of Diabetes, and
Digestive and Kidney Diseases, National Institutes of Health,
DHHS, Bethesda, Maryland, USA
ABSTRACT
A convenient synthesis of 20-deoxy-2-fluoroadenosine from commercially available
2-fluoroadenine is described. The coupling reaction of silylated 2-fluoroadenine
with phenyl 3,5-bis[O-(t-butyldimethylsilyl)]-2-deoxy-1-thio-d-erythro-pentofura-
noside gave the corresponding 2-fluoro-20-deoxyadenosine derivative (a=b ¼1:1)
in good yield. The a- and b-anomers were separated by chromatography, and
then desilylated to give compounds 1a and 1b.
Key Words: Fluorinated nucleosides; Thioglycoside coupling; Glycosylation;
Anomer separation.
INTRODUCTION
20-Deoxy-2-fluoroadenosine (F-dAdo) (1b) has received recent attention as a
prodrug for the targeted delivery of highly toxic 2-fluoroadenine (2F-Ade) to tumor
cells.[1] The strategy relies on the use of the selective expression of the E. coli purine
*Correspondence: Kenneth L. Kirk, Laboratory of Bioorganic Chemistry, National Institute
of Diabetes, and Digestive and Kidney Diseases, National Institutes of Health, DHHS,
Bethesda, MD 20892, USA; Fax: (301) 402-4182; E-mail: kennethk@intra.niddk.nih.gov.
1899
DOI: 10.1081/NCN-120025237
Copyright # 2003 by Marcel Dekker, Inc.
1525-7770 (Print); 1532-2335 (Online)
www.dekker.com

1900
Ye et al.
nucleoside phosphorylase (PNP) gene in tumor cells, and on the high substrate
activity of F-dAdo for E. coli PNP. The inactivity of this and other adenosine
analogues for mammalian PNP is the basis for tumor selectivity. Adding value to
this approach is the fact that E. coli PNP-produced 2-fluoroadenine can rapidly
diffuse across cell membranes and thus has high bystander activity, i.e., it is toxic
to tumor cells that are not proliferating or which do not have the expressed E. coli
PNP gene. In particular, the potential of increased responsiveness of solid tumors to
this approach compared to more traditional chemotherapies that are more depen-
dent on active proliferation provides a major impetus to this research. These issues
are described in greater detail in recent publications.[1,2]
Montgomery and Hewson initially prepared F-dAdo by a multi step sequence that
included fluorination of a 2,6-diaminopurine nucleoside precursor as a key step.[3]
Owing to the low yield of this procedure, Secrist and co-workers recently developed
a more convenient procedure based on free radical removal of the 20-OH group from
the more readily available 2-fluoroadenosine.[4] We are preparing a series of 2-fluoro-
20-deoxyadenosines to be used in biochemical studies of adenosine processing
enzymes. The commercial availability of 2-fluoroadenine prompted us to explore
direct glycosylation of this base with appropriate 2-deoxy-ribose partners as an
efficient route to these nucleoside derivatives. Because of the current interest in
F-dAdo as an anticancer prodrug, we report herein the successful application of this
approach to this analogue. The synthesis is short and readily scaled up.
CHEMISTRY
In preliminary experiments, using modifications of a published procedure,[5] we
found conditions for coupling thiogylcoside 2, readily prepared from 2-deoxyribose,
with 2-fluoroadenine that affords products 3 in about 59% combined yield with
1:1 a=b selectivity. Attempts to separate the anomers by chromatography were
unsuccessful. A sample of the mixture of compounds 3 was deacetylated using
5% triethylamine in methanol to give a mixture of 1a and 1b. Unfortunately we were
also unable to separate anomers 1a and 1b by chromatography on silica gel despite
repeated efforts using a variety of solvent schemes. In an effort to accentuate steric
differences imposed by the different orientation of the glycosidic bond, the mixture
of anomers 3 was deprotected and silylated in situ with TBSCl to give 4a and
4b. We were pleased to find that the silylated derivatives indeed were readily

Synthesis of 20-Deoxy-2-fluoroadenosine
1901
Scheme 1. Reagents and conditions: a) (i) HMDS, (NH4)2SO4, reflux; (ii) thioglycoside 2,
NBS, MS 4D, toluene; b) Et3N=MeOH; c) TBSCl, imidazole, DMF.
Scheme 2. Reagents and conditions: a) DowexHþ, MeOH, rt, 94%; b) TBSCl, DMAP, NEt3,
DCM=DMF, 0ꢁC to rt, 78%; c) PhSH, ZnI2, n-Bu4NI, DCM, ꢀ15ꢁC, 74%.
separated by chromatography, but the overall conversion from 3 was only 6%
(Sch. 1).
Having determined on a small scale the effectiveness of the TBS group in facil-
itating anomer separation, we saw the advantage of installing this group at the
beginning of the sequence. Thus, the TBS protected thioglycoside 7 was prepared
from the known glycoside 6[6] (Sch. 2), and this was used in the glycosylation reac-
tion. This reaction proceeded smoothly to give a 1:1 mixture of anomers 4a,b in 62%
overall yield. The mixture was separated by silica gel chromatography and removal
of the TBS group with tetraethylammonium fluoride completed the synthesis of
compounds 1a and 1b (Sch. 3). The spectral and physical data were in complete
agreement with the published data.[3,4]
EXPERIMENTAL
General Procedures
Melting points are uncorrected. 1H, 13C and 19F NMR spectra were recorded at
300.1, 75.5 and 282.2 MHz, respectively. Chemical shifts (d, ppm) of protons and
carbons are relative to TMS (0 ppm), and the fluorine shifts are relative to trifluoro-
acetic acid (ꢀ76.55 ppm). Interaction constants are presented in Hz. The solvent is
CDCl3 unless otherwise noted. Low resolution MS (LRMS) was performed with

1902
Ye et al.
Scheme 3. Reagents and conditions: a) HMDS, (NH4)2SO4, reflux; b) thiogylcoside 7, NBS,
˚
MS 4 A, toluene; 30% (4a) and 32% (4b) c) Et4NFꢂH2O, acetone, rt; 95% (1a), 87% (1b).
chemical ionization under ammonia gas. High resolution MS (HRMS) was done
with FAB ionization under xenon gas. Silica gel (0.040–0.063 mm) was used for
column chromatography. All reagents and dry solvents were purchased if not other-
wise indicated and used without further purification or drying. 2-Fluoroadenine was
purchased from Aldrich.
Methyl 2-deoxy-d-erythro-pentofuranoside (5). Dowex 50wx-100 ion-exchange
resin (100 mg) was added to a solution of 2-deoxy-D-ribose (10 g, 74.6 mmol) in
methanol (200 mL). After being stirred at room temperature for 17 h, the Dowex resin
was filtered and 2 mL of triethylamine was added to the filtrate. The solvent was
removed under reduced pressure and the residue was purified by chromatography
(EtOAc) to give a mixture of a and b anomers of 5 (10.4 g, 94%).
Methyl 3,5-Bis[O-(t-butyldimethylsilyl)]-2-deoxy-d-erythro-pentofuranoside (6). This
was prepared from 5 as described in reference.[6]
Phenyl 3,5-Bis[O-(t-butyldimethylsilyl)]-2-deoxy-1-thio-d-erythro-pentofuranoside
(7). To a solution of 6 (3.76 g, 10 mmol) in dry dichloromethane (80 mL) were
added benzenethiol (2.05 mL, 20 mmol), zinc iodide (6.38 g, 20 mmol) and tetrabutyl-
ammonium iodide (739 mg, 2 mmol). This suspension was stirred under nitrogen at
ꢀ15ꢁC for 1 h. The reaction mixture was poured into cold pH 7 phosphate buffer
and the organic layer was extracted with dichloromethane. The extract was washed
with saturated sodium chloride and dried over sodium sulfate. The solvent was
removed under reduced pressure and the residue was purified by column chromato-
graphy (hexane=ethyl acetate, 40:1) to give 3.35 g (74%) of a mixture of a and b
1
anomers of 7 as a colorless oil. H NMR (CDCl3, 300 MHz) d: 7.6–7.4 (4H, m,
20,60-SPh a and b), 7.3–7.2 (6H, m, 30,40,50-SPh a and b) 5.7–5.6 (2H, m, H-1a and
b), 4.4–4.3 (2H, m, H-4a and b), 4.1–4.0 (1H, m, H-3a or b), 4.0–3.9 (1H, m, H-
3a or b), 3.8–3.3 (4H, m, H-5a and b), 2.7–1.9 (4H, m, H-2a and b), 1.0–0.8
(36H, m, Si(C(CH3)3 a and b), 0.1–0.0 (24H, m, Si(CH3)2 a and b). 13C NMR
(CDCl3, 75 Hz) d: 137.2 and 135.2 (10-Ph) 131.5 and 130.4 (20, 60-Ph), 128.93 and

Synthesis of 20-Deoxy-2-fluoroadenosine
1903
128.86 (30, 40-Ph), 127.1 and 126.6 (50-Ph), 89.0 and 86.9 (C1), 85.8 and 85.7 (C4),
73.4 and 71.5 (C3), 63.8 and 62.5 (C5), 42.3 and 41.6 (C2), 26.15 and 26.09
(Si(C(CH3)3), 25.98 and 25.96 (Si(C(CH3)3), 18.5 and 18.2 (Si(C(CH3)3), ꢀ4.5 and
ꢀ5.2 (Si(CH3)2). Anal. Calcd for C23H42O3SSi2: C, 60.74; H, 9.31. Found: C,
61.25; H, 9.58.
2-Fluoro-9-(3,5-bis[O-(t-butyldimethylsilyl)]-2-deoxy-d-erythro-pentofuranosyl)-
adenine (4). A suspension of 2-fluoroadenine (1.53 g, 10.0 mmol) and ammonium
sulfate (1.32 g, 10.0 mmol) in hexamethyldisilazane (20 mL) was heated at vigorous
reflux under N2 until the solution became clear (5–10 h) The excess of hexamethyl-
disilazane was removed under reduced pressure. The residue and thioglycoside 7
(4.54 g, 10.0 mmol) were dissolved in 100 mL of dry toluene under N2, and then
˚
5 g of powdered molecular sieves 4 A was added. After being stirred at room tem-
perature for 30 min, the reaction mixture was cooled to 0ꢁC and NBS (1.96 g,
11.0 mmol) was added. The resulting mixture was allowed to warm to room tempera-
ture and stirred for 15 h. A small amount of silica gel was added to quench the reac-
tion. The solvent was removed under reduced pressure and the residue was purified
by chromatography on silica gel (dichloromethane=methanol, 40:1) to give 3.4 g
(68%) of a mixture of a,b anomers (1:1) as a white solid. This was rechromato-
graphed on silica gel (hexane=ethyl acetate, 7:3) to give 1.5 g (30%) of a-anomer
4a, and 1.6 g (32%) of b-anomer 4b. 4a: Mp 180–181ꢁC;1H NMR (CDCl3,
300 MHz) d: 8.31 (1H, s, H-8), 6.40 (1H, dd, J ¼ 7.5, 1.5 Hz, H-10), 5.68 (2H, br,
NH2), 4.6–4.5 (1H, m, H-40), 4.4–4.3 (1H, m, H-30), 3.73 (1H, dd, J ¼ 10.8, 3.3 Hz,
H-50), 3.60 (1H, dd, J ¼ 11.1, 5.4 Hz, H-50), 2.8–2.7 (1H, m, H-20), 2.4–2.3 (1H, m,
H-20), 0.93 (9H, s, (Si(C(CH3)3)), 0.85 (9H, s, (Si(C(CH3)3) ), 0.10 (6H, s, (Si(CH3)2)),
0.01(6H, s, (Si(CH3)2)); 13C NMR (CDCl3, 75 MHz) d: 159.3 (d, 1JCF ¼ 208 Hz, C2),
3
3
157.3 (d, JCF ¼ 20.5 Hz, C4), 151.1 (d, JCF ¼ 19.4 Hz, C6), 140.2 (C8), 117.9 (C5),
90.34 (C10), 85.32 (C40), 73.31 (C30), 63.56, (C50), 41.76, (C20), 26.09 (Si(C(CH3)3)),
25.84 (Si(C(CH3)3)), 18.51 (Si(C(CH3)3)), 18.06 (Si(C(CH3)3)), ꢀ4.68 (Si(CH3)2),
ꢀ4.77 (Si(CH3)2), ꢀ5.19 (Si(CH3)2), ꢀ5.32 (Si(CH3)2) FAB-MS m=z 498.3 [MHþ].
Anal. Calcd for C22H40FN5O3Si2: C, 53.09; H, 8.10; N, 14.07. Found: C, 53.18;
H, 8.24; N, 13.79. 4b: Mp 188–190ꢁC;1H NMR (CDCl3, 300 MHz) d: 8.31 (1H, s,
H-8), 6.36 (1H, t, J ¼ 6.3 Hz, H-10), 5.69 (2H, br, NH2), 4.6–4.5 (1H, m, H-40),
4.1–4.0 (1H, m, H-30), 3.89 (1H, dd, J ¼ 11.4, 4.2 Hz, H-50), 3.78 (1H, J ¼ dd, 11.4,
3.3 Hz, H-50), 2.7–2.5 (1H, m, H-20), 2.5–2.4 (1H, m, H-20), 0.923 (9H, s,
(Si(C(CH3)3)), 0.921 (9H, s, (Si(C(CH3)3)), 0.12 (6H, s, (Si(CH3)2)), 0.01(6H, s,
1
(Si(CH3)2)); 13C NMR (CDCl3, 75 Hz) d: 159.2 (d, JCF ¼ 209 Hz, C2), 157.3 (d,
3
3JCF ¼ 20.0 Hz, C4), 151.1 (d, JCF ¼ 19.4 Hz, C6), 139.6 (C8), 118.4 (C5), 88.16
(C10), 84.72 (C40), 72.00 (C30), 62.92, (C50), 41.43, (20), 26.15 (Si(C(CH3)3)), 26.10
(Si(C(CH3)3)), 18.62 (Si(C(CH3)3)), 18.21 (Si(C(CH3)3)), ꢀ4.46 (Si(CH3)2), ꢀ4.62
(Si(CH3)2), ꢀ5.10 (Si(CH3)2), ꢀ5.30 (Si(CH3)2) FAB-MS m=z 498.3 [MHþ]. HRMS
(FABþ) Calcd for MHþ C22H41FN5O3Si2: 498.2732; Found: 498.2723. Anal. Calcd
for C22H40FN5O3Si2: C, 53.09; H, 8.10; N, 14.07. Found: C, 52.95; H, 8.10; N, 13.72.
2-Fluoro-9-(2-deoxy-a-d-erythro-pentofuranosyl)adenine (1a). To a solution of
4a (1.02 g, 2.05 mmol) in acetone (30 mL) was added tetraethylammonium fluoride

1904
Ye et al.
hydrate (835 mg, 4.51 mmol) at room temperature. After the solution was stirred
over night the solvent was removed under reduced pressure and the residue was
purified by chromatography on silica gel (dichloromethane=methanol, 9:1) to give
527 mg (95%) of 1a as a white solid. Mp 230–232ꢁC, Lit (3) 232ꢁC 1H NMR
(DMSO-d6, 300 MHz) d: 8.35 (1H, s, H-8), 7.82 (2H, br, NH2), 6.22 (1H, dd,
J ¼ 7.8, 3.0 Hz, H-10), 5.50 (1H, dd, J ¼ 7.8, 3.0 Hz, 30-OH), 4.84 (1H, t, J ¼ 5.75 Hz,
Hz, 50-OH), 4.4–4.2 (1H, m, H-40), 4.2–4.1 (1H, m, H-30), 3.5–3.4 (2H, m, H-50), 2.8–
2.6 (1H, m, H-20), 2.31 (1H, dt, J ¼ 14.1, 3.0 Hz, H-20) 13C NMR (DMSO-d6,
1
3
75 MHz) d: 158.7 (d, JCF ¼ 202.6 Hz, C2); 157.7 (d, JCF ¼ 21.1 Hz, C4); 150.5 (d,
4
3JCF ¼ 20.5, C6); 140.0 (C8); 117.3 (d, JCF ¼ 3.98 Hz, C5); 88. 73 (C10); 83.68
(C40); 70.77 (C30), 61.74 (C50), 39.91 (C20) FAB-MS m=z 270.2 [MHþ], HRMS
(FABþ) Calcd for MHþ C10H13FN5O3: 270.1002; Found: 270.0999. Anal. Calcd
for C10H12FN5O5: C, 44.61; H, 4.49; N 26.01. Found: C, 44.31; H, 4.51, N 26.14.
2-Fluoro-9-(20-deoxy-b-d-erythro-pentofuranosyl)adenine (1b). The same proce-
dure as above using 4b instead of 4a to give 1b as a white solid. Yield: 87%. Mp
1
210ꢁC (decomp), Lit (3), >210ꢁC (indefinite) H NMR (DMSO-d6, 300 MHz) d:
8.32 (1H, s, H-8), 7.84 (2H, br, NH2), 6.23 (1H, t, J ¼ 6.6 Hz, H-10), 5.31 (1H, d,
J ¼ 4.2 Hz, 30-OH), 4.95 (1H, t, J ¼ 5.7 Hz, 50-OH), 4.4–4.3 (1H, m, H-40), 3.9–3.8
(1H, m, H-30), 3.7–3.4 (2H, m, H-50), 2.7–2.6 (1H, m, H-20), 2.3–2.2 (1H, m, H-20);
1
3
13C NMR (DMSO-d6, 75 Hz) d: 158.6 (d, JCF ¼ 179 Hz, C2), 157.7 (d, JCF
¼
3
4
21.0 Hz, C4), 150.4 (d, JCF ¼ 20.0 Hz, C6), 139.8 (C8), 117.6 (d, JCF ¼ 3.98 Hz,
C5), 87.95 (C10), 83.67 (C40), 70.77 (C30), 61.73, (C50), 39.31, (C20). FAB-MS, m=z
270 [MHþ]. HRMS (FABþ) Calcd for MHþ C10H13FN5O3: 270.1002; Found:
270.1000.
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Synthesis of 20-Deoxy-2-fluoroadenosine
1905
6. Ichikawa, Y.; Kubota, H.; Fujita, K.; Okauchi, T.; Narasaka, K. Stereoselective
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Received February 28, 2003
Accepted June 9, 2003