2444
Y. Mehellou et al. / Bioorg. Med. Chem. 18 (2010) 2439–2446
HPLC analytical investigations were conducted on a Varian Pro-
star instrument (LC work station, Varian prostar 355 LC detector)
using a Polaris C18-A 10 m column; eluant was performed using
a mobile phase consisting of a water/acetonitrile gradient.
Electrospray mass spectra were obtained using a Waters LCT
time of flight (TOF) mass spectrometer coupled to a Waters
M600 HPLC pump. Data was collected in the continuum mode over
the mass range 100–2000 amu and processed using Masslynx 4.1
software.
125 MHz): d 20.45, 20.56 (2d, J = 7.5 Hz, CH3CH), 51.69, 51.79
(CHNH), 67.23, 67.45 (2d, J = 5.0 Hz, C-500), 68.03, 68.06 (CH2Ph),
76.74 (C-20), 77.52, 77.74 (C-30), 84.18, 84.33 (2d, J = 7.5 Hz, C-40),
87.87 (C-10), 101.33 (C-5), 121.46, 121.51, 121.55, 126.21, 126.24,
129.29, 129.33, 129.40, 129.66, 129.98, 130.85, 130.86, 131.20
(Ph), 137.25 (ipso Ph), 144.25, 144.36 (C-6), 152.14, 152.26 (C-2),
166.42 (C-4), 174.75, 174.92 (2d, J = 5 Hz, C = O). MS (ES+) m/e:
584.4 (MNa+, 100%); Accurate mass: C25H28N3O10NaP required
584.1410, found 584.1418. HPLCb (H2O/CH3CN from 100/0 to 0/
100 in 20 min): tR 13.31, 13.41 min.
l
4.2. Chemistry
4.3.2. Synthesis of arabinouridine-50-[phenyl-(methoxy-
inyl)]-phosphate (6)
L-alan-
4.2.1. 1-((2R,4S,5R)-3,4-Anhydro-5-(hydroxymethyl)tetrahydro-
furan-2-yl)pyrimidin-4(1H)-one (2)
Prepared using AraU (0.20 g, 0.82 mmol), NMI (0.33 mL,
4.09 mmol), phenyl-(methoxy- -alaninyl)-phosphorochloridate
Uridine (5.00 g, 20.47 mmol) and diphenyl carbonate (4.88 g,
22.80 mmol) were suspended in DMF (40 mL). The slurry was
heated to 100 °C and sodium bicarbonate (150 mg) was then added
and the reaction mixture was heated up to 137 °C for 1.5 h. After
completion, the reaction was cooled down to room temperature,
filtered and washed with methanol (60 mL). Following drying un-
der vacuum for 3 h the title compound obtained as a white solid
in 78% yield (3.60 g). 1H NMR (DMSO-d6): d 7.83 (1H, d, J = 8.1
Hz, H-6), 6.31 (1H, m, H-10), 5.90 (1H, d, J = 8.1 Hz, H-5), 5.83
(1H, s, H-30), 5.21 (1H, s, H-40), 5.06 (1H, s, H-50), 4.37 (1H, s, 50-
OH), 4.09 (1H, s, 30-OH), 3.15–3.24 (2H, m, H-20). 13C NMR
(DMSO-d6): d 171.32 (CO), 159.79 (CO), 136.84 (C6), 108.56 (C5),
89.99 (C10), 89.18 (C20), 88.74 (C40), 74.72 (C30), 74.79 (C50).
L
(0.68 g, 2.46 mmol). The desired product was obtained as a white
solid (110.0 mg, 28%). 31P NMR (MeOD, 202 MHz): d 3.77, 3.70.
1H NMR (MeOD, 500 MHz): d 7.77, 7.75 (1H, 2d, J = 8 Hz, H-6),
7.40–7.19 (5H, m, Ph), 6.21, 6.20 (1H, 2d, J = 4.0 Hz, H-10), 5.64,
5.60 (1H, 2d, J = 8.0 Hz, H-5), 4.46–4.31 (2H, m, H-50), 4.25–4.22
(1H, m, H-20), 4.16–4.09 (2H, m, H-30, H-40), 4.04–3.98 (2H, m,
CHNH), 3.70 (3H, s, CH3O), 1.38 (1.5H, dd, J = 7.5, 0.5 Hz, CH3CH
one d.i.), 1.36 (1.5H, dd, J = 7.0, 0.5 Hz, CH3CH one d.i.). 13C NMR
(MeOD, 125 MHz): d 20.44, 20.55 (2d, J = 7.5 Hz, CH3CH), 51.52,
51.63 (CHNH), 52.87, 52.90 (CH3O), 67.23, 67.35 (2d, J = 6.25 Hz,
C-50), 76.74 (C-20), 77.55, 77.70 (C-30), 84.31, 84.38 (2d, J = 7.5 Hz,
C-40), 87.86, 87.88 (C-10), 101.29 (C-5), 121.48, 126.23, 130.85,
131.18 (Ph), 144.29, 144.37 (C-6), 152.17, 152.26 (C-2), 166.42
(C-4), 175.42, 175.59 (2d, J = 5.0 Hz, C@O). MS (ES+) m/e: 508.2
(MNa+, 100%); Accurate mass: C19H24N3O10NaP required
508.1097, found 508.1107. HPLCb (H2O/CH3CN from 100/0 to 0/
100 in 20 min): tR 10.41, 10.53 min.
4.2.2. 1-((2R,3S,4S,5R)-3,4-Dihydroxy-5-(hydroxymethyl)tetra-
hydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (3)
Compound 2 (3.50 g, 15.47 mmol) in 20 mL of 2 N HCl was
heated to 80 °C for 2 h. The reaction mixture was cooled down
and neutralized to pH ꢀ7 with sodium hydroxide. The solvent
was removed and the crude was purified by column chromatogra-
phy employing 15% methanol in DCM as an eluant. Pooling and
evaporation of the appropriate fractions gave the title compound
(2.55 g, 68% yield) as a white solid. 1H NMR (DMSO-d6): d 11.24
(1H, br s, NH), 7.64 (1H, d, H-6, J = 8.1), 5.98 (1H, m, H-10), 5.63
(1H, m, 20-OH), 5.57 (1H, d, J = 8.1 Hz, H-5), 5.53 (1H, m, 30-OH),
5.10 (1H, s, 50-OH), 4.04 (1H, m, H-20), 3.92 (1H, m, H-30), 3.75
(1H, m, H-40), 3.56–3.62 (2H, m, H-50). 13C NMR (DMSO-d6): d
163.41 (CO), 150.40 (CO), 142.28 (C-6), 99.86 (C-5), 85.05 (C-10),
84.59 (C-40), 75.28 (C-30), 75.13 (C-20), 60.67 (C-50).
4.3.3. Synthesis of arabinouridine-50-[1-naphthyl-(isopropoxy-
L-alaninyl)]-phosphate (8)
Prepared using AraU (0.20 g, 0.82 mmol), NMI (0.33 mL,
4.09 mmol), naphthyl-(isopropoxy-L-alaninyl)-phosphorochlori-
date (0.87 g, 2.46 mmol). The desired product was obtained as a
white solid (102.5 mg, 22%). 31P NMR (MeOD, 202 MHz): d 4.24,
4.17. 1H NMR (MeOD, 500 MHz): 8.19–7.37 (8H, m, Naph, H-6),
6.14 (1H, d, J = 4.0 Hz, H-10), 5.49, 5.39 (1H, 2d, J = 8.0 Hz, H-5),
4.94–4.85 (1H, m, CHCH3), 4.48–4.32 (2H, m, H-50), 4.18–4.15
(1H, m, H-20), 4.12–4.04 (2H, m, H-30, H-40) 3.99–3.92 (2H, m,
CHNH), 1.29 (3H, dd, J = 7.0, 0.5 Hz, CH3CH), 1.17–1.11 (6H, m,
(CH3)2CH). 13C NMR (MeOD, 125 MHz): d 20.47, 20.60 (2d, J = 7.5,
6.25 Hz, CH3CH), 21.91, 21.95, 21.99 ((CH3)2CH), 51.83, 51.90
(CHNH), 67.42, 67.59 (2d, J = 5.0, 6.25 Hz, C-50), 70.22, 70.25
((CH3)2CH), 76.70 (C-20), 77.61, 77.77 (C-30), 84.34, 84.41 (2d,
J = 8.75 Hz, C-40), 87.87 (C-10), 101.21 (C-5), 116.20, 116.22,
116.28, 116.31, 122.73, 122.85, 125.99, 126.58, 127.45, 127.52,
127.83, 127.91, 128.91 (Naph), 136.33 (ipso Naph), 144.22,
144.36 (C-6), 152.21 (C-2), 166.32 (C-4), 174.48, 174.72 (C@O).
MS (ES+) m/e: 586.4 (MNa+, 100%); Accurate mass: C25H30N3O10-
NaP required 586.1567, found 586.1571. HPLC (H2O/CH3CN from
100/0 to 0/100 in 20 min): tR 13.43, 13.60 min.
4.3. Standard procedure for the synthesis of phosphoramidates
AraU (1 equiv) was dissolved in 10 mL of THF/pyridine (7/3) un-
der argon. 1-Methylimidazole (NMI, 5 equiv) was added to the
reaction flask and this was followed by the addition of the appro-
priate phosphorochloridate (3 equiv) and the mixture was stirred
at room temperature for 16 h. Upon the removal of the solvent,
the crude was purified by column chromatography (CH2Cl2/CH3OH
95/5) and then preparative HPLC to give the desired product as a
white solid.
4.3.1. Synthesis of arabinouridine-50-[phenyl-(benzoxy-
inyl)]-phosphate (5)
Prepared using AraU (0.20 g, 0.82 mmol), NMI (0.33 mL,
4.09 mmol), phenyl-(benzoxy- -alaninyl)-phosphorochloridate
(0.87 g, 2.46 mmol). The desired product was obtained as a white
solid (150.0 mg, 33%). 31P NMR (MeOD, 202 MHz): d 3.76, 3.69. 1H
NMR (MeOD, 500 MHz): d 7.74 (1H, d, J = 8 Hz, H-6), 7.37–7.17
(10H, m, Ph, Ph), 6.19, 6.20 (1H, 2d, J = 4.0 Hz, H-10), 5.62, 5.56
(1H, 2d, J = 8.0 Hz, H-5), 5.17–5.12 (2H, m, CH2Ph), 4.44–4.30 (2H,
m, H-50), 4.25–4.22 (1H, m, H-20), 4.14–4.02 (3H, m, H-30, H-40,
CHNH), 1.38, 1.37 (3H, 2d, J = 8.0, 7.5 Hz, CH3CH). 13C NMR (MeOD,
L-alan-
4.3.4. Synthesis of arabinouridine-50-[phenyl-(benzoxy-
inyl)]-phosphate (4)
D-alan-
L
Prepared using AraU (0.20 g, 0.82 mmol), NMI (0.33 mL,
4.09 mmol), phenyl-(benzoxy- -alaninyl)-phosphorochloridate
D
(0.87 g, 2.46 mmol). The desired product was obtained as a white
solid (140.0 mg, 30%). 31P NMR (MeOD, 202 MHz): d 4.02, 3.50.
1H NMR (MeOD, 500 MHz): d 7.77, 7.71 (1H, 2d, J = 8.0 Hz, H-6),
7.37–7.17 (10H, m, Ph, Ph), 6.20, 6.18 (1H, 2d, J = 4.0 Hz, H-10),
5.61, 5.56 (1H, 2d, J = 8.0 Hz, H-5), 5.19–5.12 (2H, m, CH2Ph),
4.37–4.27 (2H, m, H-50), 4.23–4.22 (1H, m, H-20), 4.11–4.00 (3H,