A direct synthesis of 2-deoxy-2-fluoro-α-D-[6-3H]glucopyranosyl uridine-5′-diphosphate

Article abstract of DOI:10.1071/CH02057

The synthesis of 2-deoxy-2-fluoro-α-D[6-³H]glucopyranosyl uridine-5′-diphosphate, with the late introduction of the radiolable, has been achieved from 3,4-di-O-benzyl-2-deoxy-2-fluoro-α-D-glucosyl diphenyl phosphate, by an oxidation-reduction s

Full text of DOI:10.1071/CH02057

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Aust. J. Chem. 2002, 55, 327–329
C
H
02057
A Direct Synthesis of
2-Deoxy-2-fluoro-α-D-[6-3H]glucopyranosyl Uridine-5´-diphosphate
Robert V. StickA,B and Andrew G. WattsA
A Department of Chemistry, The University of Western Australia, 35 Stirling Highway,
Crawley, W.A. 6009, Australia.
B Author to whom correspondence should be addressed (e-mail: rvs@chem.uwa.edu.au).
The synthesis of 2-deoxy-2-fluoro-α-D-[6-3H]glucopyranosyl uridine-5´-diphosphate, with the late introduction of
the radiolabel, has been achieved from 3,4-di-O-benzyl-2-deoxy-2-fluoro-α-D-glucosyl diphenyl phosphate, by an
oxidation–reduction sequence, followed by protecting group removal and morpholidate coupling to uridine-5´-
monophosphate.
Manuscript received: 18 March 2002.
Final version: 23 May 2002.
R
.
S
i
c
k
a
d
A
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G
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a
t
5-Fluoro and 2-deoxy-2-fluoro sugars are excellent
mechanism-based inhibitors of many retaining glycoside
hydrolases, so much so that they may often be used to label
the catalytic nucleophile of the enzyme.[1,2] Naturally, some
efforts have been made to investigate the use of related
molecules, e.g. (1) and (2), for the inhibition of those
glycosyl transferases that utilize a nucleoside-diphosphate
activated sugar for the construction of the glycosidic
linkage.[3–5]
We have recently reported on our synthetic endeavours
towards (2).[6] In early inhibition experiments with (2) and
the self-glucosylating protein glycogenin, it soon became
apparent that we needed a radiolabelled version of (2), and
chose tritium. We initially prepared tritium-labelled (3)
starting from D-[6-3H]glucose—the whole procedure was
extremely unsatisfactory, generating amounts of low-level
radioactive waste at every step and yielding a product with a
very poor specific activity. We decided to rectify the
situation.
It seemed logical to develop a synthesis of (3) that
introduced the label (tritium) late in the sequence. Thus, D-
glucal was converted into the silyl ether (4); the attempted
introduction of fluorine (at C2) using Selectfluor™ was
accompanied by some loss of the silicon protecting group,
and so the silyl ether (4) was transformed into the acetate (5).
Now, treatment with Selectfluor™ gave a mixture of 2-
deoxy-2-fluoro sugars, from which the α-D-anomer (6)
fortuitously crystallized!
Treatment of the hemiacetal (6) with diphenyl
chlorophosphate gave the phosphate (7), but the attempted
removal of the acetyl group with sodium methoxide in
methanol was accompanied by some loss of the anomeric
O
O
OH
O
OH
O
R
HN
HN
HO
HO
HO
HO
OX
O
O
O
O
O
N
N
O
O
O
O
F
HO
BnO
BnO
F
O P O P O
OH OH
O P O P O
OH OH
OH OH
OH OH
(4) X = SiPh2But
(5) X = Ac
(1)
(2) R = H
(3) R = 3
H
O
H
R
OH
O
R
BnO
BnO
OAc
O
OH
O
O
HO
HO
BnO
BnO
BnO
BnO
O
O
O
P
F
F
F
F
X
O
P
X
X
O
P
OPh
O
OPh
OPh
OPh
(11) R = 3H, X = OPh
(12) R = 3H, X = O–(Bu3NH)+
(13) R = H, X = O–(Bu3NH)+
(9)
(6) X = OH
(7) X = OP(O)(OPh)2
(8) R = H
(10) R = 3
H
© CSIRO 2002
10.1071/CH02057
0004-9425/02/050327

328
R. V. Stick and A. G. Watts
75.17, 2C, CH2Ph; 75.08, d, J4,F 8.8 Hz, C4; 79.65, d, J3,F 16.0 Hz, C3;
90.43, dd, J2,P 8.0, J2,F 189 Hz, C2; 95.25, dd, J1,P 5.7, J1,F 23.2 Hz, C1;
119.93–150.19, Ph; 170.38, C=O. High-resolution mass spectrum
(FAB) m/z 637.1982 [C34H35FO9P (M + H)+• requires 637.2002].
phosphate. However, treatment of (7) with guanidine/
guanidinium nitrate in methanol gave a good yield of the
alcohol (8).[7]
Oxidation of (8) with pyridinium chlorochromate in the
presence of molecular sieves[8] smoothly gave the aldehyde
(9), and a subsequent reduction with an overly generous
amount of sodium borotritide (100 mCi), followed by
sodium borohydride, gave the alcohol (10). Treatment of
(10) with hydrogen, first in the presence of palladium-on-
carbon [to give (11)], and then platinum(IV) oxide, gave the
phosphate, isolated as the bis(tributylammonium) salt (12).
The salt (12) was converted into the labelled target (3)
according to published procedures.[6,9] The 2-deoxy-2-fluoro
sugar (3), labelled at C6 with tritium, now had an extremely
high, and useful, specific activity.
Diphenyl 3,4-Di-O-benzyl-2-deoxy-2-fluoro-α-D-glucosyl Phosphate
(8)
The phosphate (7) (305 mg, 0.48 mmol) was added to a guanidine/
guanidinium nitrate solution[7] (50 mL) and the mixture left to stir at
0°C (40 min). The solution was neutralized (Amberlite IR 120, H+
form), concentrated under vacuum and the residue purified using flash
chromatography (30% EtOAc/petrol) to give the alcohol (8) as a
colourless oil (220 mg, 79%), [α]D +64° (c, 1.1 in CHCl3) (Found: C,
64.5; H, 5.3. C32H32FO8P requires C, 64.5; H, 5.4%). 1H NMR
(500 MHz) δ 2.00, br s, OH; 3.51, dd, J5,6 2.3, J6,6 12.5 Hz, H6; 3.58,
dd, J5,6 3.1, H6; 3.63, dd, J3,4 9.1, J4,5 9.2 Hz, H4; 3.68, ddd, H5; 3.98,
dt, J2,3 9.1, J3,F 17.8 Hz, H3; 4.50, dddd, J1,2 ≈ J2,P 3.4, J2,F 48.3 Hz, H2;
4.59, 4.82, ABq, J 10.9 Hz, CH2Ph; 4.68, 4.83, ABq, J 11.0 Hz, CH2Ph;
6.04, dd, J1,P 6.4 Hz, H1; 7.18–7.45, m, 20H, Ph. 13C NMR (125.8
MHz) δ 58.90, C6; 73.96, C5; 75.57, 76.17, 2C, CH2Ph; 75.79, d, J4,F
8.9 Hz, C4; 79.87, d, J3,F 18.2 Hz, C3; 90.73, dd, J2,P 8.2, J2,F 185 Hz,
C2; 96.74, dd, J1,P 5.8, J1,F 24.0 Hz, C1; 118.25–150.27, Ph. High-
resolution mass spectrum (FAB) m/z 595.1913 [C32H33FO8P (M + H)+•
requires 595.1897].
Experimental
General experimental procedures have been given previously.[10]
Sodium boro[3H]hydride was purchased from Amersham.
Selectfluor™ was donated by Air Products and Chemicals, Inc.
6-O-Acetyl-3,4-di-O-benzyl-D-glucal (5)
Diphenyl 3,4-Di-O-benzyl-2-deoxy-2-fluoro-α-D-gluco-hexodialdo-
Acetic anhydride (5 mL) was added to 3,4-di-O-benzyl-D-glucal[11]
(6.7 g) in dry pyridine (30 mL), and the solution left to stand at room
temperature (1 h). The solution was then concentrated under vacuum
and subjected to a usual workup (EtOAc). Flash chromatography (15%
EtOAc/petrol) then gave the acetate (5) as colourless needles (7.0 g,
92%), m.p. 67–69°C (lit.[12] 71°C), [α]D +5° (lit.[12] +6.5°). The 1H
NMR spectrum was consistent with that reported.[12]
1,5-pyranosyl Phosphate (9)
Pyridinium chlorochromate (235 mg, 1.10 mmol) was added to a
vigorously stirred mixture of the alcohol (8) (130 mg, 0.22 mmol) and
3 Å molecular sieves (300 mg) in dry CH2Cl2 (10 mL), and stirring was
continued (1 h). The suspension was then filtered through Celite and
concentrated under vacuum. Rapid silica filtration (25% EtOAc/petrol)
of the residue then gave the aldehyde (9) as a colourless oil (115 mg,
1
6-O-Acetyl-3,4-di-O-benzyl-2-deoxy-2-fluoro-α-D-glucose (6)
89%) that was used without further purification. H NMR (500 MHz)
δ 3.62, dd, J3,4 9.9, J4,5 10.4 Hz, H4; 4.08, dt, J2,3 9.1, J3,F 11.7 Hz, H3;
4.09, dd, J5,6 0.6 Hz, H5; 4.57, dddd, J1,2 ≈ J2,P 3.3, J2,F 49.4 Hz, H2;
4.63, 4.81, ABq, J 10.9 Hz, CH2Ph; 4.72, 4.86, ABq, J 11.0 Hz, CH2Ph;
6.13, dd, J1,P 6.6 Hz, H1; 7.24–7.34, m, 20H, Ph; 9.39, d, H6. 13C NMR
(125.8 MHz) δ 75.13, 75.44, 2C, CH2Ph; 75.32, C5; 75.80, d, J4,F 7.0
Hz, C4; 79.45, d, J3,F 16.1 Hz, C3; 89.51, dd, J2,P 6.1, J2,F 190 Hz, C2;
94.87, dd, J1,P 5.9, J1,F 25.0 Hz, C1; 119.98–150.17, Ph; 195.11, C6.
Selectfluor™ (6 g) was added to the D-glucal (5) (4.3 g) in
dimethylformamide (10 mL) and water (20 mL), and the mixture left to
stir at room temperature (3 h). The resultant solution was concentrated
under vacuum and subjected to a usual workup (EtOAc). Flash
chromatography (25% EtOAc/petrol) then gave the hemiacetal (6) as
fine needles (2.0 g, 41%), m.p. 119–123°C (EtOH), [α]D +82° (c, 1.0
in CHCl3) (Found: C, 65.2; H, 6.2. C22H25FO6 requires C, 65.3; H,
6.2%). 1H NMR (500 MHz) δ 2.03, s, CH3; 3.52, dd, J3,4 9.2, J4,5 9.9
Hz, H4; 4.12, ddd, J5,6 2.2, 4.3 Hz, H5; 4.15, dt, J2,3 9.1, J3,F 12.3 Hz,
H3; 4.23, dd, J6,6 12.1 Hz, H6; 4.31, dd, H6; 4.51, ddd, J1,2 3.8, J2,F 49.6
Hz, H2; 4.58, 4.89, ABq, J 10.9 Hz, CH2Ph; 4.77, 4.93, ABq, J 11.2 Hz,
CH2Ph; 5.39, d, H1; 7.25–7.39, m, 10H, Ph. 13C NMR (125.8 MHz) δ
20.82, CH3; 62.80, C6; 68.64, C5; 75.11, 75.16, 2C, CH2Ph; 76.33, d,
J4,F 8.7 Hz, C4; 80.11, d, J3,F 15.8 Hz, C3; 90.45, d, J1,F 21.8 Hz, C1;
91.43, d, J2,F 190 Hz, C2; 127.86–138.05, Ph; 170.78, C=O. High-
resolution mass spectrum (fast-atom bombardment, FAB) m/z 405.1729
[C22H26FO6 (M + H)+• requires 405.1713].
Diphenyl 3,4-Di-O-benzyl-2-deoxy-2-fluoro-α-D-[6-3H]glucosyl
Phosphate (10)
A suspension of NaB[3H]4 (100 mCi) in MeOH (0.5 mL) was added to
a stirred solution of the aldehyde (9) (620 mg, 1.05 mmol) in MeOH
(5 mL), and stirring was continued (30 min). NaBH4 (19 mg,
0.50 mmol) was then added to the mixture and stirring was continued
(30 min). AcOH (0.3 mL) was added with stirring, and then the mixture
was concentrated under vacuum and subjected to a usual workup
(EtOAc). Flash chromatography (25% EtOAc/petrol) then gave the
alcohol (10) as a colourless oil (597 mg, 96%). The spectroscopic data
(1H and 13C NMR) were consistent with those observed above for (8).
Diphenyl 6-O-Acetyl-3,4-di-O-benzyl-2-deoxy-2-fluoro-α-D-glucosyl
Phosphate (7)
Bis(tributylammonium) 2-Deoxy-2-fluoro-α-D-[6-3H]glucopyranosyl
Phosphate (12)
Diphenyl chlorophosphate (0.66 mL, 3.20 mmol) was added to the
hemiacetal (6) (1.25 g, 3.11 mmol) and 4-dimethylaminopyridine
(400 mg, 3.20 mmol) in dry CH2Cl2 (10 mL) at –10°C and the solution
was left to stir at this temperature (1 h). The solution was allowed to
warm to room temperature, subjected to a usual workup (CH2Cl2) and
the residue purified using flash chromatography (20% EtOAc/petrol) to
give the phosphate (7) as a colourless oil (1.8 g, 89%), [α]D +84° (c, 1.0
in CHCl3) (Found: C, 64.1; H, 5.4. C34H34FO9P requires C, 64.2; H,
5.4%). 1H NMR (500 MHz) δ 1.95, s, CH3; 3.57, dd, J3,4 9.3, J4,5 9.7
Hz, H4; 3.78–3.94, m, H5; 4.03, dd, J5,6 2.0, J6,6 12.6 Hz, H6; 4.06, dt,
J2,3 9.2, J3,F 11.8 Hz, H3; 4.17, dd, J5,6 3.8, H6; 4.57, 4.88, ABq, J 10.9
Hz, CH2Ph; 4.60, dddd, J1,2 ≈ J2,P 3.6, J2,F 48.8 Hz, H2; 4.74, 4.86,
ABq, J 11.0 Hz, CH2Ph; 6.12, dd, J1,P 6.4 Hz, H1; 7.22–7.40, m, 20H,
Ph. 13C NMR (125.8 MHz) δ 20.55, CH3; 61.76, C6; 70.73, C5; 75.07,
Pd/C (100 mg, 10% w/w) was added to the alcohol (10) (560 mg) in
MeOH (10 mL), and the mixture was stirred vigorously under a
hydrogen atmosphere at room temperature (2 h). The mixture was
filtered through Celite, and PtO2 (40 mg) then added to the filtrate. The
mixture was again stirred vigorously under an atmosphere of hydrogen
at room temperature (overnight). The mixture was filtered through
Celite, concentrated under vacuum, and the residue taken up in water
(5 mL) and passed through a column of Dowex 50W-X8 (H+) resin into
a vigorously stirred mixture of water and excess tributylamine.
Concentration under vacuum and subsequent lyophilization gave the α-
D-phosphate (12) as
a colourless solid (542 mg, 91%). The
spectroscopic data (1H and 13C NMR) were consistent with those
reported for (13).[6]

Synthesis of 2-Deoxy-2-fluoro-α-D-[6-3H]glucopyranosyl Uridine-5´-diphosphate
329
2-Deoxy-2-fluoro-α-D-[6-3H]glucopyranosyl Uridine-5´-diphosphate
(3)
[3] C.-H. Wong, T. Hayashi, Inhibitors of Glycosyl Transferases WO
98/25940, 18th June 1998, 82 pp. (United States).
[4] T. Hayashi, B. W. Murray, R. Wang, C.-H. Wong, Bioorg. Med.
Chem. 1997, 5, 497.
The phosphate (12) (410 mg, 650 µmol) was treated with 4-
morpholine-N,N´-dicyclohexylcarboxamidinium uridine 5´-mono-
phosphomorpholidate (660 mg, 960 µmol) according to the literature
procedure[6] to give (3) as a colourless powder (268 mg, 68%). The 1H
NMR spectrum was consistent with that reported for (2).[6]
[5] R. V. Stick, D. M. G. Tilbrook, A. G. Watts, unpublished results.
[6] R. V. Stick, A. G. Watts, Monatsh. Chem. 2002, 133, 541.
[7] U. Ellervik, G. Magnusson, Tetrahedron Lett. 1997, 38, 1627.
[8] J. Herscovici, M. J. Egron, A. Kostas, J. Chem. Soc., Perkin
Trans. 1 1982, 1967.
[9] V. Wittmann, C.-H. Wong, J. Org. Chem. 1997, 62, 2144.
[10] J. C. McAuliffe, R. V. Stick, Aust. J. Chem. 1997, 50, 193.
[11] R. A. Alonso, G. D. Vite, R. E. McDevitt, B. Fraser-Reid, J. Org.
Chem. 1992, 57, 573.
References
[1] D. J. Vocadlo, G. J. Davies, R. Laine, S. G. Withers, Nature 2001,
412, 835.
[12] S. Fischer, C. H. Hamann, J. Carbohydr. Chem. 1995, 14, 327.
[2] J. D. McCarter, S. G. Withers, J. Am. Chem. Soc. 1996, 118, 241.