Synthesis and biological activity of 4′-C-hydroxymethyl-2′-fluoro-D-arabinofuranosylpurine nucleosides

Article abstract of DOI:10.1081/NCN-100105249

A series of 4′-C-hydroxymethyl-2′-fluoro-D-arabinofuranosylpurine nucleosides was prepared and evaluated for cytotoxicity. The details of a convenient synthesis of the carbohydrate precursor 4-C-hydroxymethyl-3,5-di-O-benzoyl-2-fluoro-α-D-arabinofuranosyl bromide (13) are presented. Proof of the structure and configuration at all chiral centers of the sugars and the nucleosides were obtained by proton NMR. All five target nucleosides were evaluated for cytotoxicity in human tumor cell lines. The 4′-C-hydroxymethyl clofarabine analogue (16β) showed slight cytotoxicity in CCRF-CEM leukemia cells.

Full text of DOI:10.1081/NCN-100105249

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SYNTHESIS AND BIOLOGICAL ACTIVITY OF
4′-C-HYDROXYMETHYL-2′-FLUORO- D-
ARABINOFURANOSYLPURINE NUCLEOSIDES
Anita T. Shortnacy-Fowler a , Kamal N. Tiwari a , John A. Montgomery a & John A. Secrist III b
a Southern Research Institute , P.O. Box 55305, Birmingham, AL, 35255-5305, U.S.A.
b Southern Research Institute , P.O. Box 55305, Birmingham, AL, 35255-5305, U.S.A.
Published online: 17 Aug 2006.
To cite this article: Anita T. Shortnacy-Fowler , Kamal N. Tiwari , John A. Montgomery & John A. Secrist III (2001) SYNTHESIS
AND BIOLOGICAL ACTIVITY OF 4′-C-HYDROXYMETHYL-2′-FLUORO- D-ARABINOFURANOSYLPURINE NUCLEOSIDES, Nucleosides,
Nucleotides and Nucleic Acids, 20:8, 1583-1598, DOI: 10.1081/NCN-100105249
To link to this article: http://dx.doi.org/10.1081/NCN-100105249
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NUCLEOSIDES, NUCLEOTIDES & NUCLEIC ACIDS, 20(8), 158371598 (2001)
SYNTHESIS AND BIOLOGICAL ACTIVITY
OF 40-C-HYDROXYMETHYL-20-FLUORO-D-
ARABINOFURANOSYLPURINE NUCLEOSIDES
Anita T. Shortnacy-Fowler, Kamal N. Tiwari, John A. Montgomery,
and John A. Secrist III*
Southern Research Institute, P.O. Box 55305,
Birmingham, AL 35255-5305, USA
ABSTRACT
A
series of4 0-C-hydroxymethyl-20-¯uoro-D-arabinofuranosylpurine
nucleosides was prepared and evaluated for cytotoxicity. The details of a
convenient synthesis ofthe carbohydrate precursor 4- C-hydroxymethyl-
3,5-di-O-benzoyl-2-¯uoro-a-D-arabinofuranosyl bromide (13) are pre-
sented. Proofofthe structure and con®guration at all chiral centers ofthe
sugars and the nucleosides were obtained by proton NMR. All ®ve target
nucleosides were evaluated for cytotoxicity in human tumor cell lines.
The 40-C-hydroxymethyl clofarabine analogue (16b) showed slight cyto-
toxicity in CCRF-CEM leukemia cells.
INTRODUCTION
In recent years, pursuit ofthe synthesis ofnew nucleosides has con-
tinued because oftheir demonstrated utility as anticancer and as antiviral
agents. The incorporation ofvarious groups at C-40 ofa normal nucleoside
structure has received renewed interest recently178. It has been shown by
several investigators that such a modi®cation in most cases can be tolerated
with retention or even enhancement ofthe biological activity 4,5,8. The
*Corresponding author.
1583
Copyright # 2001 by Marcel Dekker, Inc.
www.dekker.com

1584
SHORTNACY-FOWLER ET AL.
observed biological activity suggests that metabolism to the triphosphate is
likely occurring. Our laboratory has continued its investigations ofnew
nucleosides as potential anticancer agents in recent years. Fludarabine
phosphate, which was developed in our laboratory9, has been approved by
FDA for the treatment of refractory lymphocytic leukemia. Clofarabine (2-
chloro-20-¯uoro-ara-A), also developed in our laboratory10, is currently in
clinical trials. Expanding upon its structure, with a 20-¯uorine in the arabino
con®guration, we have initiated a program to examine certain other carbo-
hydrate modi®cations in the system, especially at the 40-position.
Initially, we focused our attention on the synthesis of the 40-C-hydro-
xymethyl analogue ofcloafrabine and related compounds, hoping to dis-
cover good biological activity as well as to learn more about structure
activity relationships for compounds with this modi®cation. Very few 40-C-
hydroxymethyl nucleosides have been reported in the literature, and most of
0
the work has been limited to the synthesis ofribonucleosides and 2 -
deoxynucleosides11713. 40-C-hydroxymethylthymidine has been synthe-
sized12, and derivatives ofit have been described 14716. No 40-C-hydro-
xymethyl-arabino or 20-¯uoroarabinonucleosides, however, have been
reported in the literature. Toward that goal, we have developed a route to a
4-C-hydroxymethyl-2-¯uoroarabinofuranose derivative and utilized it in the
synthesis ofcertain purine nucleosides containing that carbohydrate moiety.
All these nucleosides have been characterized, and their anomeric con®g-
urations were determined by proton NMR. Final deblocked nucleosides and
their a-anomers have been evaluated for cytotoxicity against human tumor
cell lines. We herein report the details ofa convenient synthesis ofthis 4- C-
hydroxymethyl-2-¯uoroarabinofuranose intermediate, its conversion to a
series ofpurine nucleoside analogues, and the results rfom their
testing17.
in vitro
CHEMISTRY
Carbohydrate Synthesis. Our route (Scheme 1) to this series appeared to
be best accomplished by preparing the blocked 4-C-hydroxymethyl derivative
13, which can be synthesized from a-bromo sugar 218 (easily obtained from
commercially available 1). Compound 2 was converted to 3 by the careful
addition ofNaOCH in MeOH at 0 ꢀC. When bromide displacement was
3
complete, more NaOCH3 was added with the reaction continuing at room
temperature to provide deprotected 4. Benzylation of 4 to 5 followed by
19
the selective removal ofthe primary benzyl ether
gave alcohol 6.
The 5-hydroxyl of 6 failed to oxidize to the aldehyde 7 using 1-(3-
dimethylaminopropyl)-3-ethylcarbodiimide (EDC) hydrochloride5 or
aqueous sodium periodate11. Oxidation of 6 with DMSO=acetic anhydride20
a€orded impure 7, which, when condensed with formaldehyde in base11,

40-C-HYDROXYMETHYL DERIVATIVES
1585
Scheme 1. Conditions: (a) HBr=HOAc; (b) NaOCH3=MeOH, 0 ꢀC then R.T; (c)
BnCl=KOH, THF; (d) 10% Pd=C, H2, 50 psi, 1 : 1 MeOH=HOAc, pyridine; (e) PCC, 4A sieve,
CH2Cl2; (f) 37%, HCHO, NaOH, THF=H2O; (g) 10% Pd=C, H2, 50 psi, 1 : 1 MeOH=HOAc;
(h) BzCl=pyridine; (i) TFA=H2O, 55 ꢀC; (j) Ac2O=pyridine.
produced <15% ofdiol 8. The yield of 8 was increased to 61% by a
molecular sieve catalyzed pyridinium chlorochromate oxidation21 of 6 to 7
followed by reaction with formaldehyde as above. The remaining benzyl
group was removed, and the product 9 was benzoylated to give 10. Acidic
hydrolysis ofthe methyl group of 10 provided 11 as an anomeric mixture
(2a : 1b), which was acetylated to a€ord 12 (3a : 1b)22. Treatment of 12 with
excess HBr in acetic acid gave the target 4-C-hydroxymethyl bromo sugar 13
(6a : 1b) suitable for subsequent coupling reactions.
The anomeric con®gurations ofcompounds 11, 12, and 13 were
assigned by comparison ofthe 1H NMR spectra. The a anomers had
J1,2 ˆ 0 Hz, con®rming that H-1 and H-2 were trans to each other. Similarly,
the beta anomers had J1,F ˆ 0 Hz, which con®rmed that H-1 and 2-F were in
a trans relationship23.
Nucleoside Synthesis. Our route (Scheme 2) to the purine nucleosides
began with the coupling of2,6-dichloropurine and 13 using the sodium salt
glycosylation procedure24. An 82% total yield of9-isomers 14 was obtained
after column chromatography with only minor amounts of suspected
7-isomers observed in later fractions. Separation of 14b and 14a was achieved
on small scale only, so enriched anomeric mixtures were used in ensuing steps.

1586
SHORTNACY-FOWLER ET AL.
Scheme 2. Conditions: (a) NaH, MeCN, R.T., 18 h; (b)NaOCH3=MeOH; (c) EtOH, NH3,
80 ꢀC, 16 h; (d) NaN3, EtOH, 1=2 h; (e) 5% Pd=C, H2, 1 atm, 2 : 1 EtOH=DMAc 16 h;
(f) adenosine deaminase.
Treatment of 14 with NaOCH3=MeOH removed the benzoyl groups
and replaced the 6-chloro with a methoxy group to give 15. Preparative TLC
resolved 15b and 15a, which were reacted separately with ethanolic ammonia
at 80 ꢀC to provide 2-Cl-6-NH2 analogues 16b and 16a.
Conversion of 14 to 2,6-diazido derivative 17 was carried out by treat-
ment with NaN3 in re¯uxing EtOH. When the reaction mixture was cooled,
17b crystallized and 17a remained in solution. After isolation, the separate
anomers were hydrogenated at atmospheric pressure with 5% Pd=C to give
intermediates 18b and 18a. Deprotection with NaOCH3=MeOH led to 2,6-
diamino products 19b and 19a. Deamination of 19b occurred slowly with
calfintestinal adenosine deaminase in water to produce guanosine analogue
20b. Under the same conditions, 19a appeared not to be a substrate for the
enzyme and was recovered unchanged.
The anomeric con®gurations of 14720b were determined by compar-
ison ofthe 1H NMR spectra ofthe individual anomers. All b anomers
5
showed 5-bond coupling between H-8 and 20-F, JH-8,2 F between 1.0
0
and 3.1 Hz25. In the a anomers, H-8 showed no coupling with the 20-F.
The anomeric con®gurations were further con®rmed by NOE di€erence

40-C-HYDROXYMETHYL DERIVATIVES
1587
spectroscopy. The b nucleoside 14b gave NOE's between H-8 and H-30 of
273%, and the a anomer 14a gave NOE's between H-10 and H-30 of1 72%.
0
The assignments ofH-5 and H-60 for 14720b were determined by
NOE's of1 72% between H-30 and H-50 and=or by the long range 5-bond
5
0
coupling ofone ofthe H-5 hydrogens with the 20-F, JH-5 ,2 F between 0.6
and 1.6 Hz.
0
0
BIOLOGICAL RESULTS
The 40-C-hydroxymethyl nucleosides (16a, 16b, 19a, 19b, 20b) were
examined invitro against a spectrum ofhuman tumor systems: SNB-7(CNS),
DLD-1 (colon), NCI-H23 (lung), ZR-75-1 (mammary), LOX IMVI (mela-
noma), PC-3 (prostate), CAK-1 (renal), and CCRF-CEM (leukemia). All the
compounds were found to be noncytotoxic at the highest level tested
(60 mg=mL) except 16b. This clofarabine analogue (16b) exhibited an IC50
value at 5 mg=mL (ꢁ13 mM) in CCRF-CEM cells. In this cell line, clofarabine
has an IC50 of0.05 mm10. No further toxicity for 16b was observed in any of
the other cell lines.
EXPERIMENTAL
Melting points were determined on a Mel-Temp apparatus and are
1
uncorrected. H NMR spectra were recorded on a Nicolet NT-300 NB
spectrometer operating at 300.635 MHz (1H). Chemical shifts are expressed
in parts per million down®eld from tetramethylsilane. UV absorption spectra
were determined with a Perkin-Elmer Lambda 9 spectrometer by dissolving
each compound in methanol or water and diluting 10-fold with 0.1 N HCl,
pH 7 bu€er, or 0.1 N NaOH. Numbers in parentheses are extinction coef-
®cients (Â10À 3), sh ˆ shoulder. Microanalyses were performed by the
Spectroscopic and Analytical Laboratory ofSouthern Research Institute.
Where solvents were noted as part ofthe elemental analyses, they were seen
1
in the H NMR spectra in the proper amounts. Mass spectra were recorded
on a Varian=MAT 311A double-focusing mass spectrometer in the fast atom
bombardment (FAB) mode. HPLC analyses were carried out at a ¯ow rate of
1 ml=min on a Hewlett-Packard 1100 series liquid chromatograph with a
Phenomenex Sphereclone 5 m ODS(1) column (4.6 Â 250 mm) and UV mon-
itoring (254 nm). Flash chromatographic separations were performed using
2307400 mesh silica gel from E. Merck. TLC was done on Analtech pre-
coated (250 mm) silica gel (GF) plates. Compounds isolated as syrups were
single spots on TLC or two spots for anomeric mixtures. No other impurities
were seen in the NMR spectra ofthese compounds.
Methyl 3,5-Di-O-benzyl-2-¯uoro-b-D-arabinofuranoside (5). To a solution of
bromo sugar 218 (5 g, 11.8 mmol) in dry MeOH (50 mL) at 0 ꢀC was added a

1588
SHORTNACY-FOWLER ET AL.
0.5 M solution ofNaOCH in MeOH (23.64 mL) dropwise in a period of
3
30 min. Additional 0.5 M NaOCH3 in MeOH (50 mL) was added, and the
solution was stirred at room temperature for 2 h. The solution was rendered
‡
neutral with Dowex 50W- X8 (H ) ion exchange resin, and the resin was
®ltered o€ with MeOH washing. The ®ltrates were combined and evaporated
to dryness to a€ord crude 4, which was a single spot on TLC (Rf 0.50, 9 : 1
CHCl3=MeOH). This material was dissolved in dry THF (100 mL), and
powdered KOH (2.64 g, 47 mmol) and benzyl chloride (3.45 mL; 30 mmol)
were added to it. The suspension was re¯uxed overnight, then ®ltered.
Collected solids were washed with THF, and the ®ltrate and washings were
combined and evaporated to dryness. This residue was puri®ed on a silica gel
column using 91 : 9 cyclohexane=EtOAc as eluent to obtain pure 5 (3.2 g,
79%) as an oily wax: TLC 3 : 1 cyclohexane=EtOAc, Rf 0.55; MS m=z 347
‡
1
(M‡H) ; H NMR (CDCl3) d7.2577.35 (m, 10H, aromatic H0s), 4.9675.15
(m, 1H, H-2), 4.9574.97 (m, 1H, H-1), 4.5674.72 (m, 4H, PhCH2), 4.1274.26
(m, 2H, H-3, H-4), 3.5473.58 (m, 2H, H-5), 3.40 (s, 3H, CH3).
Methyl 3-O-Benzyl-2-deoxy-2-¯uoro-b-D-arabinofuranoside (6). To
a
solution of 5 (3.41 g, 9.86 mmol) in 1 : 1 MeOH=HOAc (33 mL) was added
pyridine (34 mL) and 10% palladium on charcoal (463 mg). The mixture was
hydrogenated at 50 psi for 22 h. The catalyst was removed by ®ltration and
washed with several portions ofMeOH. The combined ®ltrate and washes
were evaporated to a residue, which was dissolved in EtOAc (65 mL), washed
with saturated NaHCO3 (20 mL) and water (2 Â 20 mL), dried (MgSO4), and
evaporated to give 6 (1.76 g, 70%) as a syrup: TLC 1 : 1 hexane=EtOAc, Rf
‡ 1
0.60; MS m=z 257 (M‡H) ; H NMR (CDCl3) d 7.2677.36 (m, 5H, aromatic
H's), 4.9775.15 (m, 1H, H-2), 4.9374.95 (m, 1H, H-1), 4.5674.77 (m, 2H,
PhCH2), 4.3774.47 (m, 1H, H-3), 4.1174.15 (m, 1H, H-4), 3.6173.80 (m, 2H,
H-5), 3.50 (s, 3H, CH3), 2.19. (dd, 1H, OH-5, J ˆ 4.5 Hz and J ˆ 7.8 Hz).
Methyl 3-O-Benzyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-b-D-arabinofurano-
side (8). To a solution of6 (3.08 g, 12.0 mmol) in anhydrous CH 2Cl2
(72 mL) under argon was added 4A molecular sieve powder (10.9 g). After
being stirred at room temperature for 20 min, the mixture was treated in one
portion with solid pyridinium chlorochromate (5.28 g, 24.0 mmol) at which
time the reaction became very dark and thickened. After 3 h, the vigorously
stirred mixture was ®ltered through a dry silica gel pad (59 g, 2307400 mesh),
which was then washed with multiple portions of3 : 1 EtOAc =hexane
(500 mL total volume). The ®ltrate plus washes were evaporated to give
‡
crude 7 (2.75 g, 90% as a yellow oil): MS m=z 261 (M‡Li) . This unstable
aldehyde was dissolved in THF (30 mL), diluted with water (30 mL), and
chilled to 075 ꢀC. To this cloudy solution was added dropwise aqueous 37%
formaldehyde (4.5 mL) followed by 2N NaOH (12 mL). The cooling bath was
removed, and the reaction was stirred at room temperature 16 h, neutralized

40-C-HYDROXYMETHYL DERIVATIVES
1589
to pH 6 with glacial acetic acid, and evaporated. The resulting residue was
partitioned between CH2Cl2 and water. The aqueous layer was extracted
twice with more CH2Cl2, and the combined organic layers were washed once
with water, dried (MgSO4), and evaporated to provide impure 8 (3.4 g). This
material was puri®ed by ¯ash chromatography on silica gel (58 g) with 1 : 1
hexane=EtOAc as solvent to a€ord pure 8 (2.11 g, 61% from 6) as a colorless
‡
1
syrup: TLC 1 : 1 hexane=EtOAc, Rf 0.25; MS m=z 293 (M‡Li) ; H NMR
(CDCl3) d7.3077.40 (m, 5H, aromatic H's), 5.16 (ddd, 1H, H-2, J1,2 ˆ 4.7 Hz,
J2,3 ˆ 7.4 Hz, J2,F ˆ 53.4 Hz), 4.97 (dd, 1H, H-1, J1,F ˆ 1.3 Hz, J1,2 ˆ 4.7 Hz),
5
4.83 (dd, 1H, PhCH2, JH,F ˆ 1.3 Hz, J ˆ 11.6 Hz), 4.63 (d, 1H, PhCH2,
J ˆ 11.6 Hz), 4.53 (dd, 1H, H-3, J2,3 ˆ 7.4 Hz, J3,F ˆ 18.0 Hz), 3.70 (dd, 1H,
H-6b, J6a,6b ˆ 12.2 Hz, J6b,OH ˆ 7.0 Hz), 3.65 (dd, 1H-6a, J6a,6b ˆ 12.2 Hz,
J6a,OH ˆ 7.4 Hz), 3.57 (d, 2H, H-5, J ˆ 5.7 Hz), 3.51 (s, 3H, CH3), 2.38 (dd,
1H, OH-6, J6a,OH ˆ 7.4 Hz, J6b,OH ˆ 7.0 Hz), 2.28 (t, 1H, OH-5,
J5,OH ˆ 5.7 Hz).
Methyl 2-Deoxy-2-¯uoro-4-C-hydroxymethyl-b-D-arabinofuranoside (9). A
solution of 8 (1.82 g, 6.36 mmol) in 1 : 1 MeOH=acetic acid (70 mL)
containing 10% palladium on charcoal (352 mg) was hydrogenated at
50 psi for 22 h. More catalyst (160 mg) was added, and hydrogenation was
resumed for 24 h. The catalyst was collected and washed with several
portions ofMeOH. The combined ®ltrates were evaporated, and the
resulting oil was coevaporated with toluene to remove acetic acid. The
residue was puri®ed by ¯ash chromatography on silica gel (30 g) with a
gradient from 1 : 1 hexane=EtOAc to 100% EtOAc to yield essentially pure 9
(962 mg, 77%) as a syrup: TLC 3 : 1 EtOAc=hexane, Rf 0.20; MS m=z 197
‡
(M‡H) . This material was used directly below.
Methyl 3,5,6-Tri-O-benzoyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-b-D-arabino-
furanoside (10). To an ice-cold solution of 9 (962 mg, 4.91 mmol) in
anhydrous pyridine (25 mL) was added dropwise benzoyl chloride (2.9 mL,
24.5 mmol). The solution was allowed to warm to room temperature where it
was kept for 24 h and then poured into a stirred ice=water mixture (300 mL).
CHCl3 (100 mL) was added, and after 15 min, the layers were separated. The
aqueous layer was extracted with more CHCl3 (3 Â 100 mL). The combined
organic layers were washed with water (100 mL), dried (MgSO4), and
evaporated to an oil, which was coevaporated twice with toluene. The
residue was puri®ed by ¯ash chromatography on silica gel (75 g) with
gradient elution from 5 : 1 to 3 : 1 hexane=EtOAc to provide 10 (2.37 g,
95%) as a colorless glass: TLC 3 : 1 hexane=EtOAc, Rf 0.50; MS m=z 509
‡
(M‡H) ; 1H NMR (CDCl3) d 7.9078.05 (m, 6H, o-H ofPh), 7.3477.54 (m,
9H, m- and p-H ofPh), 6.11 (dd, 1H, H-3, J 2,3 ˆ 7.8 Hz, J3,F ˆ 17.5 Hz), 5.39
(ddd, 1H, H-2, J1,2 ˆ 4.5 Hz, J2,3 ˆ 7.7 Hz, J2,F ˆ 53.1 Hz), 5.12 (dd, 1H, H-1,
J1,F ˆ 1.2 Hz, J1,2 ˆ 4.5 Hz), 4.75 (s, 2H, H-6), 4.59 (dd, 1H, H-5a,

1590
SHORTNACY-FOWLER ET AL.
5
J5a,5b ˆ 12.0 Hz, JH,F ˆ 0.6 Hz), 4.51 (d, 1H, H-5b, J5a,5b ˆ 12.0 Hz), 3.51 (s,
3H, CH3).
3,5,6-Tri-O-benzoyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-D-arabinofuranoside
(11). A solution of 10 (2.34 g, 4.61 mmol) in 9 : 1 tri¯uoroacetic acid=water
(23 mL) was maintained at 55 ꢀC for 44 h then cooled to room temperature
and diluted with CH2Cl2 (25 mL). This solution was added slowly to a stirred
mixture ofice and saturated NaHCO 3 (250 mL). More CH2Cl2, ice, and solid
NaHCO3 were added during the addition to control foaming and to keep the
pH at ꢁ7. The layers were separated, and the aqueous layer was extracted
with more CH2Cl2 (2 Â 100 mL). The combined organic layers were washed
with water (2 Â 150 mL), dried (MgSO4), and evaporated to a white foam
(2.19 g). This material was ¯ash chromatographed on silica gel (50 g) with
gradient elution from 3 : 1 to 2 : 1 hexane=EtOAc to give pure 11 (1.8 g, 79%,
a : b, 2 : 1) as a colorless glass: TLC 3 : 1 hexane=EtOAc, Rf 0.33; MS m=z 495
‡
1
(M‡H) ; H NMR (CDCl3) d 7.8978.10 (m, 6H, o-H ofPh), 7.52 77.60
(m, 3H, p-H ofPh), 7.32 77.45 (m, 6H, m-H ofPh), 6.05 (dd, 1H, H-3 b,
J2,3 ˆ 6.0 Hz, J3,F ˆ 17.3 Hz), 5.97 (d, 1H, H-3a, J3,F ˆ 20.8 Hz, J2,3 ˆ 0 Hz),
5.71 (br d, 1H, H-1a, J1,F ˆ 9.6 Hz, J1,2 ˆ 0 Hz), 5.6375.68 (br m, 1H, H-1b),
5.30 (ddd, 1H, H-2b, J1,2 ˆ 4.0 Hz, J2,3 ˆ 6.0 Hz, J2,F ˆ 52.5 Hz), 5.20 (d, 1H,
H-2a, J1,2 ˆ J2,3 ˆ 0 Hz, J2,F ˆ 49.1 Hz), 5.18 (d, 1H, H-5ba, J5a,5b ˆ 11.9 Hz),
5.00 (d, 1H, H-5bb, J5a,5b ˆ 11.9 Hz), 4.4374.75 (m, 6H, H-5aa, H-5ab, H-6a
and H-6b), 3.6673.64 (br m, 1H, OH-1a and OH-1b).
1-O-Acetyl-3,5,6-tri-O-benzoyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-a- and -b-
D-arabinofuranoside (12). A solution of 11 (1.55 g, 3.14 mmol) in anhydrous
pyridine (15 mL) at 075 ꢀC was treated dropwise with acetic anhydride
(0.90 mL, 9.54 mmol). After 10 min, the solution was allowed to warm to
room temperature where it was held for 22 h. The solution was evaporated in
vacuo to a sti€ syrup, which was puri®ed by ¯ash chromatography on
silica gel (50 g) with 3 : 1 hexane=EtOAc as solvent. Compound 12 (1.68 g,
100%, a : b, 3 : 1) was isolated as a colorless glass: TLC 3 : 1 hexane=EtOAc,
‡
‡
Rf a 0.43 and Rf b 0.48; MS m=z 554 (M‡NH4) , 477 (M-OAc) ; 1H NMR
(CDCl3) d 7.8978.10 (m, 6H, o-H ofPh), 7.50 77.60 (m, 3H, p-H ofPh),
7.3477.45 (m, 6H, m-H ofPh), 6.54 (d, 1H, H-1 b, J1,F ˆ 0, J1,2 ˆ 4.6 Hz), 6.51
(d, 1H, H-1a, J1,F ˆ 11.1 Hz, J1,2 ˆ 0), 6.11 (dd, 1H, H-3a, J2,3 ˆ 7.6 Hz,
J3,F ˆ 17.6 Hz), 6.03 (d, 1H, H-3b, J3,F ˆ 19.0 Hz), 5.54 (ddd, 1H, H-2b,
J1,2 ˆ 4.6 Hz, J2,3 ˆ 7.6 Hz, J2,F ˆ 52.9 Hz), 5.27 (d, 1H, H-2a, J2,F
ˆ
49.0 Hz), 4.5674.85 (m, 8H, H-5a, H-5b, H-6a, and H-6b), 2.16 (s, 3H,
CH3-a), 2.09 (s, 3H, CH3-b).
3,5,6-Tri-O-benzoyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-a and b-D-arabino-
furanosyl bromide (13). To an ice-cold solution of 12 (1.68 g, 3.13 mmol) in
anhydrous CH2Cl2 (20 mL) was added dropwise over 15 min 30% HBr in

40-C-HYDROXYMETHYL DERIVATIVES
1591
acetic acid (16 mL). The clear yellow solution in a tightly sealed ¯ask was
allowed to warm to room temperature, stirred 15 h, and evaporated invacuo.
The resulting residue was partitioned between CH2Cl2 (50 mL) and ice-cold
saturated NaHCO3 (50 mL). The layers were separated, and the organic layer
was washed with ice water (2 Â 50 mL), dried (MgSO4), and evaporated. This
unstable 13 (a : b, 6 : 1) was held under vacuum until used in the reaction
below: TLC 3 : 1 hexane=EtOAc, Rf a 0.62 and Rf b 0.52; 1H NMR (CDCl3)
d 7.3278.10 (m, 15H, Ph H's), 6.69 (d, 1H, H-1b, J1,F ˆ 0, J1,2 ˆ 4.8 Hz), 6.62
(d, 1H, H-1a, J1,F ˆ 13.4 Hz, J1,2 ˆ 0), 6.26 (d, 1H, H-3b, J2,3 ˆ 7.8 Hz, J3,F
15.7 Hz), 6.07 (d, 1H, H-3a, J3,F ˆ 18.0 Hz), 5.62 (d, 1H, H-2a, J2,F
ˆ
ˆ
49.6 Hz), 5.48 (ddd, 1H, H-2b, J1,2 ˆ 4.8 Hz, J2,3 ˆ 7.8 Hz, J2,F ˆ 54.6 Hz),
4.5874.95 (m, 8H, H-5a, H-5b, H-6a, and H-6b).
2,6-Dichloro-9-(3,5,6-tri-O-benzoyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-b- and -
a-D-arabinofuranosyl)-9H-purine (14b and 14a). A suspension of2,6-
dichloropurine (650 mg, 3.44 mmol) in anhydrous MeCN (20 mL) under
argon at room temperature was treated portionwise over 15 min with NaH
(60% dispersion in mineral oil, 158 mg, 3.95 mmol). After an additional
20 min, a solution of 13 (1.74 g, 3.13 mmoles) in MeCN (8 mL) was added
dropwise over 20 min, and the mixture was stirred at room temperature 18 h.
Insoluble solids were collected and washed with CH2Cl2. The combined ®ltrate
and washes were evaporated to an orange foam, which was ¯ash
chromatographed on silica gel (50 g). Elution with 3 : 1 hexane=isopropyl
acetate a€orded mixed products, 14b and 14a as white foams (1.70 g, 82%),
which were used in subsequent reactions. Small samples ofenriched fractions
were further puri®ed by preparative TLC (Analtech GF, 10Â 20 cm, 1,000 m)
with multiple development in 100 : 1 CHCl3=MeOH to provide material for
analysis.
Compound 14b: TLC 100 : 1 CHCl3=MeOH, Rf 0.45; MS m=z 671
‡ 1
(M‡Li) ; H NMR (CDCl3) d 8.44 (d, 1H, H-8, J8,F ˆ 3.1 Hz), 7.9578.12 (m,
0
6H, o-H ofPh), 7.4377.65 (m, 9H, m-H and p-H ofPh), 6.87 (dd, 1H, H-1 ,
J1 ,2 ˆ 3.3 Hz, J1 ,F ˆ 19.8 Hz), 6.27 (dd, 1H, H-30, J2 ,3 ˆ 1.5 Hz, J3 ,F
ˆ
0
0
0
0
0
0
17.4 Hz), 5.45 (ddd, 1H, H-20, J2 ,3 ˆ 1.5 Hz, J1 ,2 ˆ 3.3 Hz, J2 ,F ˆ 50.5 Hz),
0
0
0
0
0
5
5.02 (dd, 1H, H-50b, J5 ,F ˆ 0.7 Hz, J5 a,5 b ˆ 12.0 Hz), 4.93 (d, 1H, H-60b,
0
0
0
J6 a,6 b ˆ 11.9 Hz), 4.75 (bd, 1H, H-50a, J5 a,5 b ˆ 12.0 Hz), 4.60 (d, 1H, H-60a,
0
0
0
0
0
0
J6 a,6 b ˆ 11.9 Hz).
Compound 14a: TLC 100 : 1 CHCl3=MeOH, Rf 0.40; MS m=z 671
‡
1
(M‡Li) ; H NMR (CDCl3) d 8.44 (s, 1H, H-8); 7.8478.12 (m, 6H, o-H of
0
Ph), 7.3677.61 (m, 9H, m-H and p-H ofPh), 6.54 (dd, 1H, H-1
,
J1 ,2 ˆ 3.6 Hz, J1 ,F ˆ 14.4 Hz), 6.24 (dd, 1H, H-30, J2 ,3 ˆ 3.4 Hz, J3 ,F
ˆ
0
0
0
0
0
0
23.6 Hz), 6.19 (ddd, 1H, H-20, J1 ,2 ˆ 3.6 Hz, J2 ,3 ˆ 3.4 Hz, J2 ,F ˆ 43.9 Hz),
0
0
0
0
0
5
4.97 (dd, 1H, H-50b, J5 a,5 b ˆ 12.0 Hz, J5 ,F ˆ 1.0 Hz), 4.88 (d, 1H, H-60b,
0
0
0
J6 a,6 b ˆ 12.0 Hz), 4.70 (d, 1H, H-50a, J5 a,5 b ˆ 12.0 Hz), 4.67 (bd, 1H, H-60a,
0
0
0
0
0
0
J6 a,6 b ˆ 12.0 Hz).

1592
SHORTNACY-FOWLER ET AL.
2-Chloro-6-methoxy-9-(2-deoxy-2-¯uoro-4-C-hydroxymethyl-b-D-arabinofurano-
syl)-9H-purine (15b). A solution of 14 (5b : 1a) (172 mg, 0.26 mmol) in MeOH
(10 mL) at room temperature was treated with 0.5 N NaOCH3 in MeOH
(0.52 mL) and stirred 4 h. Strong cation exchange resin (hydrogen form) was
added to neutralize, and the mixture was stirred 10min. The resin was
collected and washed with several portions ofMeOH. The combined ®ltrate
and washes were evaporated, and the residue was puri®ed by preparative TLC
(Analtech GF, 20Â 20cm, 2,000 m) developed three times in 9 : 1
CHCl3=MeOH. A MeOH extract ofthe product band gave taefr
evaporation essentially pure 15b (56 mg, 75%) as a gummy solid: TLC 9 : 1
CHCl3=MeOH, Rf 0.38; HPLC 98%, tR ˆ 8.6 min, 4 : 1 H2O=MeCN; MS m=z
‡
1
355 (M‡Li) ; H NMR (DMSO-d6) d 8.61 (d, 1H, H-8, J8,F ˆ 1.0 Hz), 6.56
(dd, 1H, H-10, J1 ,F ˆ 5.6 Hz, J1 ,2 ˆ 6.3 Hz), 6.03 (bs, 1H, OH), 5.55 (ddd, 1H,
0
0
0
H-20, J1 ,2 ˆ 6.3 Hz, J2 ,3 ˆ 6.5 Hz, J2 ,F ˆ 56.7 Hz), 5.22 (bs, 1H, OH), 5.04 (bs,
0
0
0
0
0
1H, OH), 4.65 (dd, 1H, H-30, J2 ,3 ˆ 6.5 Hz, J3 ,F ˆ 26.3 Hz), 4.11 (s, 3H, CH3),
0
0
0
3.64 (bd, 1H, H-50b, J5 a,5 b ˆ 11.5 Hz), 3.54 (bs, 2H, H-60), 3.41 (dd, 1H, H-50a,
0
0
5
0
0
0
J5 a,F ˆ 1.6 Hz, J5 a,5 b ˆ 11.5 Hz).
2-Chloro-6-methoxy-9-(2-deoxy-2-¯uoro-4-C-hydroxymethyl-a-D-arabinofurano-
syl)-9H-purine (15a). Compound 14 (7a : 1b) (62 mg, 0.09 mmol) was reacted
with 0.5 N NaOCH3 (0.18 mL) as described above for preparing 15b to
provide 15a (23 mg, 82%) as a gum: TLC 9 : 1 CHCl3=MeOH, Rf 0.44; HPLC
‡
1
98%, tR ˆ 9.5 min, 4 : 1 H2O=MeCN; MS m=z 355 (M‡Li) ; H NMR
(DMSO-d6)d 8.75 (s, 1H, H-8), 6.23 (dd, 1H, H-10, J1 ,2 ˆ 5.6 Hz,
0
0
J1 ,F ˆ 13.9 Hz), 5.70 (ddd, 1H, H-20, J1 ,2 ˆ 5.6 Hz, J2 ,3 ˆ 6.6 Hz,
0
0
0
0
0
J2 ,F ˆ 55.5 Hz), 5.15 (bs, 1H, OH), 4.61 (dd, 1H, H-30, J2 ,3 ˆ 6.6 Hz,
0
0
0
J3 ,F ˆ 21.2 Hz), 4.12 (s, 3H, CH3), 3.68 (d, 1H, H-50b, J5 a,5 b ˆ 11.8 Hz),
0
0
0
5
3.44 (dd, 1H, H-50a, J5 a,F ˆ 1.3 Hz, J5 a,5 b ˆ 11.8 Hz), 3.43 (bs, 2H, H-60).
0
0
0
2-Chloro-9-(2-deoxy-2-¯uoro-4-C-hydroxymethyl-b-D-arabinofuranosyl)-9H-
purin-6-amine (16b). A solution of 15b (62 mg, 0.18 mmol) in 60 mL of
ethanolic ammonia (saturated at 5 ꢀC) in a glass-lined stainless steel bomb
was kept at 80 ꢀC for 16 h and then evaporated. The residue was puri®ed by
preparative TLC (Analtech GF, 20 Â 20 cm, 2,000 m) with two developments
in 5 : 1 CHCl3=MeOH‡1% concd NH4OH. A warm MeOH extract ofthe
product band provided after evaporation a residue that failed to crystallize.
An acetone solution ofthis material was allowed to evaporate to a glass,
which was scratched to a powder a€ording pure 16b (50 mg, 75%): TLC 5 : 1
CHCl3=MeOH‡1% concd NH4OH, Rf 0.33; HPLC 97%, tR ˆ 5.4 min, 4 : 1
‡
NH4H2PO4 (0.01 M, pH 5.1)=MeOH; MS m=z 334 (M‡H) ; UV lmax pH 1,
1
264 (14.1), pH 7, 263 (14.6), pH 13, 264 (14.9); H NMR (DMSO-d6)d
5
8.32 (d, 1H, H-8, J8,F ˆ 1.0 Hz), 7.87 (bs, 2H, NH2), 6.46 (dd, 1H, H-10,
J1 ,2 ˆ 6.3 Hz, J1 ,F ˆ 5.9 Hz), 5.92 (d, 1H, 30-OH, J3 ,3 -OH ˆ 5.4 Hz),
0
0
0
0
0
5.50 (dt, 1H, H-20, J1 ,2 ˆ J2 ,3 ˆ 6.3 Hz, J2 ,F ˆ 56.7 Hz), 5.13 (t, 1H, 60-OH,
0
0
0
0
0

40-C-HYDROXYMETHYL DERIVATIVES
1593
J6 ,6 -OH ˆ 5.6 Hz), 4.98 (dd, 1H, 50-OH, J5 b,5 -OH ˆ 5.7 Hz, J5 a,5 -OH ˆ 5.4 Hz)
0
0
0
0
0
0
4.64 (ddd, 1H, H-30, J2 ,3 ˆ 6.3 Hz, J3 ,F ˆ 26.1 Hz, J3 ,3 -OH ˆ 5.4 Hz), 3.63
0
0
0
0
0
(dd, 1H, H-50b, J5 a,5 b ˆ 11.5 Hz, J5 b,5 -OH ˆ 5.7 Hz), 3.52 (d, 2H, H-60, J6 ,6 -
0
0
0
0
0
0
5
ˆ 5.6 Hz), 3.39 (ddd, 1H, H-50a, J5 a,5 b ˆ 11.5 Hz, J5 a, F ˆ 1.0 Hz, J5 a,5 -
0
0
0
0
0
OH
OH
ˆ 5.6 Hz). Anal. Calcd. for C11H13ClFN5O4 Á 0.5 acetone Á 0.5 H2O: C,
40.39; H, 4.61; N, 18.84. Found: C, 40.61; H, 4.45; N, 18.94.
2-Chloro-9-(2-deoxy-2-¯uoro-4-C-hydroxymethyl-a-D-arabinofuranosyl)-9H-
purin-6-amine (16a). Compound 15a (32 mg, 0.09 mmol) was reacted with
ethanolic ammonia as described for preparing 16b to give 16a (20 mg,
64%) as a white solid: TLC 5 : 1 CHCl3=MeOH‡1% concd NH4OH, Rf
0.40; HPLC 100%, tR ˆ 6.6 min, 4 : 1 NH4H2PO4 (0.01 M, pH 5.1)=MeOH;
‡
MS m=z 334 (M‡H) ; UV lmax pH 1, 264 (15.0), pH 7, 264 (15.2), pH 13
1
264 (15.2); H NMR (DMSO-d6)d 8.48 (s, 1H, H-8), 7.92 (bs, 2H, NH2),
6.12 (dd, 1H, H-10, J1 ,2 ˆ 5.8 Hz, J1 ,F ˆ 13.6 Hz), 5.94 (d, 1H, 30-OH,
0
0
0
J3 ,3 -OH ˆ 5.6 Hz), 5.64 (ddd, 1H, H-20, J1 ,2 ˆ 5.8 Hz, J2 ,3 ˆ 6.5 Hz,
0
0
0
0
0
0
J2 ,F ˆ 55.5 Hz), 5.06 (dd, 1H, 50-OH, J5 b,5 -OH ˆ 5.6 Hz, J5 a,5 -
0
0
0
0
0
ˆ 6.0 Hz), 4.96 (t, 1H, 60-OH, J6 ,6 -OH ˆ 5.4 Hz), 4.59 (ddd, 1H, H-30,
0
0
OH
J2 ,3 ˆ 6.5 Hz, J3 ,F ˆ 21.1 Hz, J3 ,3 -OH ˆ 5.6 Hz), 3.67 (dd, 1H, H-50b, J5 b,5 -
0
0
0
0
0
0
0
ˆ 5.6 Hz, J5 a,5 b ˆ 11.8 Hz), 3.4173.47 (m, 3H, H-50a and H-60). Anal.
0
0
OH
Calcd. for C11H13ClFN5O4 Á 0.4 H2O: C, 38.76; H, 4.08; N, 20.54. Found:
C, 38.79; H, 4.00; N, 20.34.
2,6-Diazido-9-(3,5,6-tri-O-benzoyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-b-and-
a-D-arabinofuranosyl)-9H-purine (17b and 17a). To a solution of 14 (9b : 1a)
(1.27 g, 1.91 mmol) in warm EtOH (50 mL) was added dropwise with stirring
a solution ofNaN (289 mg, 4.40 mmol) in H2O (1.3 mL). The mixture was
3
re¯uxed 0.5 h (TLC, 2 : 1 hexane=EtOAc), cooled to room temperature, and
then chilled in ice for 2 h. The solid present was collected, washed with cold
EtOH, and air dried before being dissolved in CHCl3. This solution was
washed once with H2O, dried (MgSO4), and evaporated to give 17b (902 mg,
77%) as a white solid ofsucient purity for the next step. The EtOH ®ltrate
from the reaction mixture contained the alpha anomer and other minor
impurities. The solvent was evaporated, and the residue was processed as
described above to provide crude 17a (456 mg). A small sample was puri®ed
for analysis by preparative TLC as reported for 14a.
Compound 17b: TLC 100 : 1 CHCl3=MeOH, Rf 0.45; MS m=z 679
‡
1
5
(M‡H) ; H NMR (CDCl3) d 8.19 (d, 1H, H-8, J8,F ˆ 2.8 Hz), 7.9478.08
(m, 6H, o-H ofPh), 7.38 77.64 (m, 9H, m-H and p-H ofPh), 6.78 (dd, 1H,
H-10, J1 ,2 ˆ 3.5 Hz, J1 -F ˆ 19.4 Hz), 6.29 (dd, 1H, H-30, J2 ,3 ˆ 1.9 Hz,
0
0
0
0
0
J3 ,F ˆ 17.1 Hz), 5.43 (ddd, 1H, H-20, J2 ,3 ˆ 1.9 Hz, J1 ,2 ˆ 3.5 Hz,
0
0
0
0
0
5
J2 ,F ˆ 50.8 Hz), 4.98 (dd, 1H, H-50b, J5 b,F ˆ 0.6 Hz, J5 a,5 b ˆ 12.1 Hz), 4.90
0
0
0
0
(d, 1H, H-60b, J6 a,6 b ˆ 12.0 Hz), 4.74 (d, 1H, H-50a, J5 a,5 b ˆ 12.1 Hz), 4.59
0
0
0
0
(d, 1H, H-60a, J6 a,6 b ˆ 12.0 Hz).
0
0

1594
SHORTNACY-FOWLER ET AL.
Compound 17a: TLC 100 : 1 CHCL3=MeOH, Rf 0.40; MS m=z 679
‡
1
(M‡H) ; H NMR (CDCl3) d 8.18 (s, 1H, H-8), 7.8678.11 (m, 6H, o-H of
0
Ph), 7.3477.60 (m, 9H, m-H and p-H ofPh), 6.46 (dd, 1H, H-1
,
J1 ,2 ˆ 4.0 Hz, J1 ,F ˆ 14.2 Hz), 6.25 (ddd, 1H, H-20, J1 ,2 ˆ 4.0 Hz,
0
0
0
0
0
J2 ,3 ˆ 3.7 Hz, J2 ,F ˆ 51.3 Hz), 6.23 (dd, 1H, H-30, J2 -3 ˆ 3.7 Hz,
0
0
0
0
0
5
J3 ,F ˆ 16.6 Hz), 4.95 (dd, 1H, H-50b, J5 b,F ˆ 0.7 Hz, J5 a,5 b ˆ 11.8 Hz), 4.85
0
0
0
0
(d, 1H, H-60b, J6 a,6 b ˆ 11.9 Hz), 4.69 (d, 1H, H-60a, J6 a,6 b ˆ 11.9 Hz), 4.68
0
0
0
0
(dd, 1H, H-50a, J5 a,5 b ˆ 11.8 Hz).
0
0
9-(3,5,6-Tri-O-benzoyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-b-D-arabinofurano-
syl)-9H-purine-2,6-diamine (18b). A solution of 17b (926 mg, 1.37 mmol) in 2 : 1
EtOH=N,N-dimethylacetamide (150 mL, warmed to dissolve) was treated
with 5% palladium on carbon (120 mg) and hydrogenated for 16 h at room
temperature and atmospheric pressure. The catalyst was removed by ®ltration
and washed thoroughly with EtOH, then CHCL3. The ®ltrate plus washes
were evaporated invacuo to a yellow glass. This material was puri®ed by ¯ash
chromatography on silica gel (35 g) with 95 : 5 CHCL3=MeOH as eluting
solvent. The resulting foam solidi®ed to a white powder upon sonication with
ether to a€ord pure 18b (696 mg, 81%): TLC 95 : 5 CHCl3=MeOH‡1%
‡
concd NH4OH, Rf 0.48; MS m=z 627 (M‡H) ; 1H NMR (CDCl3) d
5
7.9578.05 (m, 6H, o-H ofPh), 7.79 (d, 1H, H-8, J8,F ˆ 2.9 Hz), 7.5477.59
0
(m, 3H, p-H ofPh), 7.36 77.45 (m, 6H, m-H ofPh), 6.59 (dd, 1H, H-1 ,
J1 ,2 ˆ 3.8 Hz, J1 ,F ˆ 18.7 Hz), 6.44 (dd, 1H, H-30, J2 ,3 ˆ 2.4 Hz,
0
0
0
0
0
J3 ,F ˆ 16.9 Hz), 5.43 (ddd, 1H, H-20, J2 ,3 ˆ 2.4 Hz, J1 ,2
ˆ
3.8 Hz,
0
0
0
0
0
J2 ,F ˆ 51.0 Hz), 5.37 (bs, 2H, 6-NH2), 4.93 (bd, 1H, H-50b,
0
J5 a,5 b ˆ 11.6 Hz), 4.89 (dd, 1H, H-60b, J6 a,6 b ˆ 11.8 Hz), 4.86 (dd, 1H, H-
0
0
0
0
50a, J5 a,5 b ˆ 11.6 Hz, J5 a,F ˆ 0.8 Hz), 4.80 (bs, 2H, 2-NH2), 4.64 (d, 1H, H-60a,
0
0
0
0
0
J6 a,6 b ˆ 11.8 Hz).
9-(3,5,6-Tri-O-benzoyl-2-deoxy-2-¯uoro-4-C-hydroxymethyl-a-D-arabinofurano-
syl)-9H-purine-2,6-diamine (18a). Compound 17a (388 mg, 0.57 mmol) was
hydrogenolyzed with 5% Pd=C (50 mg) as described for preparing 18b to
provide 18a (305 mg, 85%) as a foam: TLC 95 : 5 CHCL3=MeOH‡1% concd
‡
NH4OH, Rf 0.52; MS m=z 627 (M‡H) ; 1H NMR (CDCL3 d 7.8378.10 (m,
6H, o-H ofPh), 7.72 (s, 1H, H-8), 7.30 77.59 (m, 9H, m-H and p-H ofPh),
6.45 (dt, 1H, H-20, J1 ,2 ˆ J2 ,3 ˆ 4.5 Hz, J2 ,F ˆ 51.6 Hz), 6.25 (dd, 1H, H-10,
0
0
0
0
0
J1 ,2 ˆ 4.5 Hz, J1 ,F ˆ 10.0 Hz), 6.19 (dd, 1H, H-30, J2 ,3 ˆ 4.5 Hz,
0
0
0
0
0
J3 ,F ˆ 12.8 Hz), 5.41 (bs, 2H, 6-NH2), 5.10 (d, 1H, H-60b, J6 a,6 b ˆ 12.0 Hz),
0
0
0
4.91 (d, 1H, H-50b, J5 a,5 b ˆ 12.0 Hz), 4.80 (bs, 2H, 2-NH2), 4.77 (d, 1H, H-60a,
0
0
J6 a,6 b ˆ 12.0 Hz), 4.75 (d, 1H, H-50a, J5 a,5 b ˆ 12.0 Hz).
0
0
0
0
9-(2-Deoxy-2-¯uoro-4-C-hydroxymethyl-b-D-arabinofuranosyl)-9H-purine-2,6-
diamine (19b). A suspension of 18b (691 mg, 1.10 mmol) in MeOH (50 mL)
at room temperature was treated in one portion with 0.5 N NaOCH3 in

40-C-HYDROXYMETHYL DERIVATIVES
1595
MeOH (1.10 mL). A clear solution was observed after 5 min, and the
reaction was stirred an additional 2 h. After being neutralized with glacial
acetic acid, the reaction was evaporated. The resulting residue was puri®ed
by preparative TLC (Analtech GF, 20 Â 20 cm, 2,000 m) with two
developments in 30 : 10 : 1 CHCl3=MeOH=NH4OH. A hot MeOH extract
ofthe product band was evaporated to dryness. From this material, pure
19b (327 mg, 86%) was obtained as a gelatinous white solid from EtOH:
mp 142 ꢀC; TLC 30 : 10 : 1 CHCL3=MeOH=NH4OH, Rf 0.35; HPLC 100%,
tR ˆ 5.9 min, 3 : 1 NH4H2PO4 (0.01 M, pH 5.1)MeOH; MS m=z 321
‡
(M‡Li) ; UV lmax pH 1, 252 (12.2), 290 (10.6), pH 7, 255 (10.1), 279
(10.7), pH 13, 255 (9.9), 279 (10.7); 1H NMR (DMSO-d6) d 7.86 (d, 1H, H-8,
5
J8,F ˆ 1.3 Hz), 6.75 (bs, 2H, 6-NH2), 6.37 (dd, 1H, H-10, J1 ,2 ˆ 6.2 Hz,
0
0
J1 ,F ˆ 7.6 Hz), 5.86 (d, 1H, 30-OH, J3 -3 -OH ˆ 5.3 Hz), 5.81 (bs, 2H, 2-NH2),
0
0
0
5.37 (ddd, 1H, H-20, J2 ,3 ˆ 5.9 Hz, J1 ,2 ˆ 6.2 Hz, J2 ,F ˆ 56.8 Hz), 5.23 (bt,
0
0
0
0
0
1H, 60-OH, J6 ,6 -OH ˆ 5.0 Hz), 4.93 (dd, 1H, 50-OH, J5 a,5 -OH ˆ 5.3 Hz, J5 b,5 -
0
0
0
0
0
0
ˆ 5.6 Hz), 4.60 (ddd, 1H, H-30, J3 ,3 -OH ˆ 5.3 Hz, J2 ,3 ˆ 5.9 Hz,
0
0
0
0
OH
J3 ,F ˆ 25.4 Hz), 3.62 (dd, 1H, H-50b, J5 b,5 -OH ˆ 5.6 Hz, J5 a,5 b ˆ 11.5 Hz),
0
0
0
0
0
3.47 (d, 2H, H-60, J6 ,6 -OH ˆ 5.0 Hz), 3.35 (dd, 1H, H-50a, J5 a,F ˆ 0.8 Hz,
0
0
0
0
0
0
0
J5 a,5 -OH ˆ 5.3 Hz, J5 a,5 b ˆ 11.5 Hz). Anal. Calcd. for C11H15FN6O4 Á 0.75
H2O Á 0.25 CH3CO2H: C, 40.29; H, 5.15; N, 24.52. Found: C, 40.20; H,
4.98; N, 24.52.
9-(2-Deoxy-2-¯uoro-4-C-hydroxymethyl-a-D-arabinofuranosyl)-9H-purine-2,6-
diamine (19a). Compound 18b (305 mg, 0.49 mmol) was treated as described
for the preparation of 19b from 18b. Pure 19a (115 mg, 75%) crystallized
from EtOH: mp 2057206 ꢀC; TLC 30 : 10 : 1 CHCl3=MeOH=NH4OH, Rf
0.48; HPLC 100%, tR ˆ 7.7 min, 3 : 1 NH4H2PO4 (0.01 M, pH 5.1)=MeOH;
‡
MS m=z 315 (M‡H) ; UV lmax pH 1, 252 (12.1), 290 (10.3), pH 7, 255 (9.9),
280 (10.3), pH 13, 256 (10.2), 279 (10.8); 1H NMR (DMSO-d6) d 8.02 (s, 1H,
H-8), 6.83 (bs, 2H, 6-NH2), 6.00 (dd, 1H, H-10, J1 ,2 ˆ 6.2 Hz,
0
0
J1 ,F ˆ 13.8 Hz), 5.93 (d, 1H, 30-OH, J ˆ 5.7 Hz), 5.82 (bs, 2H, 2-NH2),
0
5.58 (ddd, 1H, H-20, J1 ,2 ˆ 6.2 Hz, J2 ,3 ˆ 6.6 Hz, J2 -F ˆ 55.8 Hz), 5.21
0
0
0
0
0
(dd, 1H, 50-OH, J5 a,5 -OH ˆ 4.9 Hz, J5 b,5 -OH ˆ 6.3 Hz), 5.05 (t, 1H, 60-OH,
0
0
0
0
J6 a,6 -OH ˆ 5.7 Hz), 4.55 (ddd, 1H, H-30, J3 ,3 -OH ˆ 5.7 Hz, J2 ,3 ˆ 6.6 Hz,
0
0
0
0
0
0
J3 ,F ˆ 21.1 Hz), 3.64 (dd, 1H, H-50b, J5 b,5 -OH ˆ 6.3 Hz, J5 a,5 b ˆ 11.8 Hz),
0
0
0
0
0
5
3.44 (dd, 1H, H-50a, J55 a,F ˆ 1.2 Hz, J5 a,5 -OH ˆ 4.9 Hz, J5 a,5 b ˆ 11.8 Hz),
3.3773.43 (m, 2H, H-60). Anal. Calcd. for C11H15FN6O4: C, 42.04; H, 4.81;
N, 26.74. Found: C, 41.96; H, 4.76; N, 26.65.
0
0
0
0
0
2-Amino-9-(2-deoxy-2-¯uoro-4-C-hydroxymethyl-b-D-arabinofuranosyl)-1,9-
dihydro-6H-purin-6-one (20b). A solution of 19b (56 mg, 0.16 mmol) in H2O
(5 mL) at room temperature was treated with adenosine deaminase
(7.9 mg, ꢁ12 units, Type II: crude powder, Sigma). After 94 h, the
reaction mixture was diluted with H2O (5 mL), and this solution was

1596
SHORTNACY-FOWLER ET AL.
applied to a strong cation exchange column (1 Â 12 cm) hydrogen form
(AG 50W-X4, 1007200 mesh, Bio-Rad) equilibrated with H2O. Gradient
elution from 07100% 0.25 N NH4OH with UV monitoring at 254 nm
provided product containing fractions that were combined and
evaporated. The resulting residue was coevaporated with EtOH and then
triturated with acetone to give 20b (45 mg, 84%) as a white solid: mp
2627264 ꢀC; TLC 9 : 2 MeCN=1N NH4OH, Rf 0.40; HPLC 99%,
tR ˆ 3.8 min, 3 : 1 NH4H2PO4 (0.01 M, pH 5.1)=MeOH; MS m=z 316
‡
(M‡H) ; UV lmax pH 1, 256 (12.2), 282 (sh), pH 7, 252 (13.3), 275 (sh),
1
pH 13, 263 (11.1); H NMR (DMSO-d6) d 10.25 (bs, 1H, 1-NH), 7.87 (d,
1H, H-8, 5J8,F ˆ 1.3 Hz), 6.52 (bs, 2H, NH2), 6.31 (dd, 1H, H-10, J1 ,2 ˆ 6.0
0
0
Hz, J1 ,F ˆ 7.2 Hz), 5.88 (bs, 1H, 30-OH), 5.37 (ddd, 1H, H-20, J1 ,2 ˆ 6.0 Hz,
0
0
0
J2 ,3 ˆ 6.3 Hz, J2 ,F ˆ 56.8 Hz), 5.14 (bs, 1H, 60-OH), 4.94 (bs,1H, 50-OH),
0
0
0
4.55 (dd, 1H, H-30, J2 ,3 ˆ 6.3 Hz, J3 ,F ˆ 25.6 Hz), 3.61 (bd, 1H, H-50b,
0
0
0
J5 a,5 b ˆ 11.5 Hz), 3.47 (bs, 2H, H-60), 3.34 (bdd, 1H, H-50a, J5 a,F ˆ 1.3 Hz,
0
0
0
0
0
J5 a,5 b ˆ 11.5 Hz). Anal. Calcd. for C11H14FN5O5 Á 0.10 EtOH Á 0.50 H2O:
C, 40.90; H, 4.78; N, 21.29. Found: C, 41.01; H, 4.75; N, 21.08.
ACKNOWLEDGMENT
This investigation was supported by the National Cancer Institute,
National Institutes ofHealth (P01-CA34200). The authors are indebted to
Dr. J. M. Riordan for recording and interpreting NMR spectra, to Mr. M.
Richardson for mass-spectral data, to Ms. J. Bearden for UV and elemental
analyses, and to Ms. S. Campbell for HPLC analyses. We gratefully
acknowledge Dr. W. Waud, Dr. W. Parker, and Ms. D. Adamson for
cytotoxicity data. The authors wish to thank Ms. Stephanie Brannon for
assistance preparing the manuscript.
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Received November 14, 2000
Accepted February 23, 2001