3248 J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 17
Manfredini et al.
compound 7 (free base) as a gray solid: yield 27%; mp 87-93
°C (MeOH/Et2O); UV (MeOH) λmax 273 nm (ꢀ 7300), λmin 253
1H, Hb-allyl); 5.20-4.80 (m, 3H, Hc-allyl and H5); 4.24-4.10
(m, 1H, H3′); 4.08-3.90 (m, 6H, Ha-allyl, H2′, H4′, H5′, H5”);
31P NMR (DMSO-d6) δ -7.87 (d, J p,p ) 20.33 Hz, 1P, PR);
-15.36 (d, J p,p ) 20.2 Hz, 1P, Pâ); MALDI MS (M + Na)+
466.2 Da.
1
nm (ꢀ 5500); H NMR (DMSO-d6) δ 7.58 (d, J ) 7.4 Hz, 1H,
H6); 7.15 (m, 2H, NH2); 6.14 (d, J ) 5.0 Hz, 1H, H1′); 5.75-
5.50 (m, 3H, Hb-allyl, H5 and OH); 5.15-4.95 (m, 3H, Hc-
allyl and OH); 4.15-3.50 (m, 7H, H2′, H3′, H4′, H5′, H5” and
Ha-allyl); MALDI MS (M)+ 283.8 Da. Anal. (C12H17N3O5) C,
H, N.
3′,5′-Bis-O-ter t-bu tyl-d im eth ylsilyl-a r a A (8). Compound
8 was prepared following and adapting the procedure reported
by Baker et al.10 Ara-A (250 mg, 0.94 mmol) was dissolved in
anhydrous DMF (5 mL) under positive argon pressure, and
freshly distilled Et3N (0.65 mL, 4.7 mmol) and TBDMS-Cl (531
mg, 3.52 mmol) were then added. After 72 h at 60 °C (TLC:
CH2Cl2/MeOH, 9:1) the mixture was diluted with EtOAc (10
mL) and washed with H2O (10 mL). The organic layer was
dried and evaporated, and the crude residue was purified by
silica gel column chromatography (CH2Cl2/MeOH, 9.5:0.5) to
give 348 mg of 8: yield 74%; mp 175-178.10
3′,5′-Bis-O-ter t-bu tyld im eth ylsilyl-2′-O-a llyl-a r a A (9).
Compound 8 (174 mg, 0.35 mmol) was dissolved in THF (6
mL), and NaH 60% (35 mg, 0.88 mmol) was added under
vigorous stirring; after 15 min allyl bromide (75 µL, 0.88 mmol)
was added and the mixture was stirred under positive argon
pressure at room temperature for 16 h (TLC: EtOAc/hexane,
8:2). When the reaction was complete, the solvent was evapo-
rated and the residue was dissolved in CH2Cl2 (10 mL), washed
with aqueous saturated NH4Cl solution (2 × 10 mL), dried,
and evaporated. The resulting solid was purified by silica gel
column chromatography (EtOAc/hexane, 6:4) to give 61 mg of
9: yield 32%; white foam; 1H NMR (CDCl3) δ 8.34 (s, 1H, H2);
8.20 (s, 1H, H8); 6.52 (d, J ) 5.1 Hz, 1H, H1′); 6.00 (sbr, 2H,
NH2); 5.62-5.45 (m, 1H, Hb-allyl); 5.38-4.92 (m, 2H, Hc-allyl);
4.52-4.49 (m, 1H, H3′); 4.05-3.65 (m, 6H, H2′, H4′, H5′, H5”
and Ha-allyl); 0.91 and 0.93 (s, 18H, 2 × tBut-Si); 0.14, 0.09,
0.07 and 0.02 (s, 12H, 2 × Me2-Si).
2′-O-Allyl-3′,5′-bis-O-ter t-bu tyldim eth ylsilyl-a r a U (13a ).
Compound 4 (200 mg, 0.7 mmol) was dissolved in anhydrous
DMF (5 mL); freshly distilled Et3N (0.66 mL, 5.5 mmol) and
TBDMS-Cl (351 mg, 3.85 mmol) were then added, and the
mixture was heated at 60 °C under vigorous stirring and under
positive argon pressure for 20 h (TLC: CH2Cl2/MeOH, 9.5:
0.5). The solvent was evaporated; the residue was dissolved
in CH2Cl2 (10 mL), washed with saturated NaHCO3 (1 × 10
mL) and brine (1 × 10 mL), dried, filtered, and evaporated.
The resulting solid was purified by silica gel column chroma-
tography (EtOAc/hexane, 3:7) to give 183 mg of compound
13a : yield 51%; white foam; 1H NMR (CDCl3) δ 8.50 (sbr, 1H,
NH); 7.80 (d, J ) 8 Hz, 1H, H6); 6.37 (d, J ) 4.6 Hz, 1H, H1′);
5.94-5.60 (m, 2H, Hb-allyl and H5); 5.40 (dd, J ) 6.6 Hz and
J ) 1 Hz, 2H, Hc-allyl); 4.38-4.00 (m, 3H, H2′, H3′ and H4′);
3.95-3.60 (m, 4H, Ha-allyl, H5′ and H5”); 0.93 and 0.91 (s,
18H, 2 × tBut-Si); 0.12 and 0.1 and 0.08 and 0.03 (s, 12H, 2 ×
Me2-Si); MALDI MS (M + Na)+ 535.9 Da; (M + K)+ 552.0
Da.
2′-O-Allyl-3′-O-ter t-bu tyld im eth ylsilyl-a r a U (14a ). Com-
pound 13a (676 mg, 1.32 mmol) was dissolved in 80% acetic
acid (10 mL), and the solution was stirred at 60 °C for 6 h
(TLC: EtOAc/hexane, 1:1). The mixture was then coevaporated
with EtOH (4 × 10 mL), and the residue was dissolved in (CH2-
Cl2), washed with aqueous saturated NaHCO3 solution (1 ×
10 mL) and brine (1 × 10 mL), dried, and evaporated. The
resulting crude solid was purified by silica gel column chro-
matography (EtOAc/hexane, linear gradient from 3:7 to 8:2)
to give 283 mg of 14a : yield 54%; white foam; 1H NMR (CDCl3)
δ 8.25 (sbr, 1H, NH); 7.70 (d, J ) 6 Hz, 1H, H6); 6.10 (d, J )
4 Hz, 1H, H1′); 5.80-5.50 (m, 2H, H5 and Hb-allyl); 5.10 (dd,
J ) 1 Hz and J ) 6.6 Hz, 2H, Hc-allyl); 4.90 (sbr, 1H, OH);
4.30-4.10 (m, 3H, H2′, H3′ and H4′); 3.85-3.75 (m, 2H, H5′
and H5”); 3.78-3.45 (m, 2H, Ha-allyl); 0.80 (s, 9H, tBut-Si);
0.03 and 0.018 (s, 6H, Me2-Si); MALDI MS (M + Na)+ 422.1
Da; (M + K)+ 438.6 Da.
2′-O -Ally l-3′-O -t er t -b u t y ld im e t h y ls ily l-5′-O -t o s y l-
a r a U (15a ). Compound 14a (283 mg, 0.71 mmol) was dissolved
in anhydrous CH2Cl2 (6 mL), and 4-dimethylaminopyridine
(DMAP) (174 mg, 1.42 mmol) was added under positive argon
pressure. Tosyl chloride (203 mg, 1 mmol) was dissolved in
freshly distilled CH2Cl2 (2 mL), and the solution obtained was
added dropwise at the reaction mixture under vigorous stirring
at 0 °C. After 20 h at 4 °C and 30 min at room temperature,
TLC (EtOAc/hexane, 4:6) indicated complete reaction; the
solvent was evaporated, and the residue was purified by silica
gel column chromatography (EtOAc/hexane, 3:7); 330 mg of
compound 15a was obtained: yield 84%; white foam; 1H NMR
(CDCl3) δ 8.84 (sbr, 1H, NH); 7.75 (d, J ) 8.4 Hz, 2H, Ph);
7.30 (d, J ) 8.4 Hz, 2H, Ph); 7.25 (d, J ) 6 Hz, 1H, H6); 6.15
(d, J ) 3.8 Hz, 1H, H1′); 5.58-5.50 (m, 2H, Hb-allyl and H5);
5.06 (dd, J ) 1 Hz and J ) 6.6 Hz, 2H, Hc-allyl); 4.40-4.10
(m, 3H, H2′, H3′ and H4′); 3.94-3.88 (m, 2H, H5′ and H5”);
3.84-3.77 (m, 2H, Ha-allyl); 2.39 (s, 3H, Me-Ph); 0.80 (s, 9H,
tBut-Si); 0.03 and 0.01 (s, 6H, Me2-Si); MALDI MS (M + Na)+
576.2 Da; (M + K)+ 592.5 Da.
2′-O-Allyl-a r a A (10). The protected compound 9 (57 mg,
0.11 mmol) was dissolved under positive argon pressure in dry
MeOH (2 mL), and NH4F (47 mg, 1.27 mmol) was added. The
mixture was heated at reflux conditions until complete conver-
sion to deprotected compound 10 (TLC: CH2Cl2/MeOH, 9:1).
After evaporation, the crude residue was purified by silica gel
column chromatography (CH2Cl2/MeOH, 9.5:0.5) to provide 29
mg of 10: yield 82%; white solid; mp 219-224 °C (MeOH/
1
Et2O); UV (MeOH) λmax 260 nm (ꢀ 12500); H NMR (DMSO-
d6) δ 8.22 (s, 1H, H2); 8.13 (s, 1H, H8); 7.27 (sbr, 2H, NH2);
6.39 (d, J ) 5.7 Hz, 1H, H1′); 5.70-5.45 (m, 2H, Hb-allyl and
OH); 5.15-4.70 (m, 3H, Hc-allyl and OH); 4.5-4.3 (m, 1H,
H3′); 4.30-3.55 (m, 6H, H2′, H4′, H5′, H5” and Ha-allyl);
MALDI MS (M)+ 307.6 Da. Anal. (C13H17N5O4) C, H, N.
2′-O-Allyl-a r a C 5′-Dip h osp h a te (12). Compound 7 (80 mg,
0.28 mmol) and Proton Sponge (150 mg, 0.7 mmol) were
dissolved in anhydrous trimethyl phosphate (1.4 mL), and
freshly distilled POCl3 (52 µL, 0.56 mmol) was added dropwise
under positive argon pressure at 0 °C. After 4 h (TLC: i-PrOH/
NH4OH/H2O, 11:7:2) a solution of 0.5 M tri-n-butylammonium
orthophosphate (n-But3NH+H2PO4-) (2.8 mL, 1.4 mmol) and
tributylamine (0.28 mL) was added to the stirred suspension
followed by a 0.2 M solution of triethylammonium bicarbonate
(TEAB) (pH 7.5, 15 mL). The solvent was then evaporated,
maintaining bath temperature at about 40-45 °C, and the
residue was purified by a DEAE Sephadex A-25 (2 × 30 cm)
column with TEAB (linear gradient from 0.01 to 1 M), pH 7.5,
at a flow rate of 33 mL/h over 10 h. Tetra-n-butylammonium
cation was exchanged for sodium by passing the solution
through a Dowex AG 50W-X2 column (50-100 mesh, Na+
form) and eluting with 10 column volumes of deionized water.
After freeze-drying of appropriate fractions, the solid mass
obtained was further purified by HPLC with CH3CN/H2O
(linear gradient from 1:9 to 6:4) to give, on freeze-drying, 19
mg of compound 12: retention time ) 5.90; yield 13%; white
solid; mp > 300 °C (MeOH/Et2O); UV (H2O) λmax 278 nm (ꢀ
10200) and 242 nm (ꢀ 9500); 1H NMR (CD3OD) δ 7.95 (d, J )
7.6 Hz, 1H, H6); 6.27 (d, J ) 4.6 Hz, 1H, H1′); 5.90-5.65 (m,
2′-O-Allyl-5′-O-tosyl-a r a U (16a ). Compound 15a (322 mg,
0.58 mmol) was dissolved in CH3CN (15 mL), and 2% HF/H2O
(3 × 15 mL) was added at room temperature every 12 h (TLC:
CH2Cl2/MeOH, 9.5:0.5). The mixture was neutralized with
saturated NH4HCO3, and the solvent was evaporated to give,
on freeze-drying, a crude residue, which was dissolved in a
CH2Cl2/MeOH (9:1) solution, filtered, and purified by column
chromatography (CH2Cl2/MeOH, linear gradient from 9.8:0.2
1
to 9.5:0.5) to give 226 mg of 16a : yield 89%; white foam; H
NMR (CDCl3) δ 8.80 (sbr, 1H, NH); 7.73 (d, J ) 8.4 Hz, 2H,
OTs); 7.32 (d, J ) 8.4 Hz, 2H, OTs); 7.20 (d, J ) 6 Hz, 1H,
H6); 6.10 (d, J ) 3.8 Hz, 1H, H1′); 5.55-5.50 (m, 3H, Hb-allyl,
H5 and OH); 5.05 (dd, J ) 1 Hz and J ) 6.6 Hz, 2H, Hc-allyl);