S. Yajima et al. / Bioorg. Med. Chem. 14 (2006) 2799–2809
2807
d 4.88 (s, 2H), d 4.69 (m, 1H), d 3.72–2.89 (m, majority),
d 2.55 (m, 1H), d 1.88 (m, 2H), d 1.76 (m, 2H), d 1.69 (m,
2H), MALDI-TOF MS, found M+H+, M+Na+, M+K+
(calcd M+H+, M+Na+, M+K+) 642.10, 664.16, 680.17
(641.69, 663.67, 679.78).
Rf = 0.275 (solvent 4), 1H NMR (D2O, 300 MHz) d
8.10 (s, 1H), d 5.41 (d, 1H), d 4.79 (s, 2H), d
4.05 (t, 1H), d 3.67–3.02 (m, majority), d 2.83 (m,
2H), d 2.33 (s, 3H), d 2.27 (s, 3H), d 1.87 (s,
3H), d 1.26 (s, 6H).
4.29. MethylaN[(Guanine-9-yl)acetyl]-NG-(2,2,4,6,7-pen-
tamethyl-dihydrobenzofuran-5-sulfonyl)arginate (Gb-R(Pbf)-
OMe, 25)
4.32. 60-N-[aN[(Guanine-9-yl)acetyl]-D- or L-arginyl]neamine
(Gb-(D)R-neamine, 28D and Gb-(L)R-neamine, 28L)
Compound 27 (10.6 mg, 13.1 lmol) was dissolved in
TFA (0.8 ml), cooled to 0 ꢁC, and stirred for 2 h.
The reaction mixture was poured into cold diethyl
ether. The precipitated was collected by centrifuge
and washed with diethyl ether 7 times. The crude
compound was dissolved in 10% acetic acid solution
and purified by HPLC. Gb-(D)R-Neamine, 28D.
Yield: 6.9 mg, 10.3 mmol, 79.2%, 1H NMR (D2O,
300 MHz) d 7.81 (s, 1H), d 4.88 (d, 1H), d 4.82 (s,
2H), d 3.94 (t, 1H), d 3.74–2.88 (m, majority), d
2.22 (m, 1H), d 1.73–1.59 (m, 2H), MALDI-TOF
MS, found M+H+, M+Na+, M+K+ (calcd M+H+,
M+Na+,M+K+), 670.1, 692.2, 708.2 (669.9, 692.9,
708.9).
Compound 4 (0.093 g, 0.454 mmol) and H-Arg(Pbf)-
OMe HCl (0.100 g, 0.227 mmol) were suspended in
DMF (1.0 ml),
a
solution of PyBrop (0.275 g,
0.590 mmol, in DMF 1.0 ml) added at 0 ꢁC. Subse-
quently, a DMF solution (1.0 ml) of DIEA (0.16 ml,
0.908 mmol) was added slowly and stirred for over-
night at room temperature. The reaction mixture was
added 20 ml of CHCl3 and 80 ml of water, extracted
three times, concentrated, and dried. The product was
obtained as a yellow oil. Yield: 0.226 g. Rf = 0.675 (sol-
1
vent 3), H NMR (DMSO-d6, 300 MHz) d 10.54 (br s,
1H), d 8.68 (d, 1H), d 7.57 (s, 1H), d 6.39 (s, 2H), d
4.67 (s, 2H), d 3.60 (s, 3H), d 3.32 (q, 1H), d 2.93 (s,
2H) 2.50 (s, 3H), d 2.46 (s, 3H), d 1.99 (s, 3H), d
1.39 (s, 6H).
Gb-(L)R-Neamine, 30L was obtained in the same
manner
as
Gb-(D)R-neamine.
Yield:
6.0 mg,
4.30. N[(Guanine-9-yl)acetyl]-NG-(2,2,4,6,7-pentameth-
9.0 lmol, 69.2% from 13, 1H NMR (D2O,
300 MHz) d 7.87 (s, 1H), d 5.49 (d, 1H), d 4.85
(s, 2H), d 4.16 (t, 1H), d 3.79–3.08 (m, majority),
d 2.27 (m, 1H), d 1.70–1.57 (m, 2H), MALDI-
TOF MS, found M+H+, M+Na+ (calcd M+H+,
M+Na+), 670.1, 692.0 (669.9, 692.9).
a
yl-dihydrobenzofuran-5-sulfonyl)arginate (Gb-R(Pbf)-
OH, 26)
Compound 25 (0.226 g, 0.357 mmol) was dissolved in
methanol (5 ml), 1 M NaOH soln was added to the solu-
tion of 12 and stirred for 1 h at room temperature. The
pH of the solution was adjusted to 3 by HCl soln and
stored in a refrigerator overnight. Resultant precipitate
was collected and washed with water, and dried in vac-
uum in the presence of P2O5. Yield: 0.064, 0.103 mmol,
28.9%. Rf = 0.6 (solvent 3), 1H NMR (DMSO-d6,
300 MHz) d 10.58 (br s, 1H), d 8.49 (d, 1H), d 7.60 (s,
1H), d 6.44 (s, 2H), d 4.68 (s, 2H), d 3.01 (q, 1H), d
2.95 (s, 2H) 2.48 (s, 3H), d 2.42 (s, 3H), d 2.00 (s, 3H),
d 1.40 (s, 6H).
4.33. tert-Butyl N[(thymine-1-yl)acetyl]-eN-(benzyloxy-
carbonyl) lysinate (Tb-K(Cbz)-OH, 29)
a
Thymine acetic acid, 4 (0.182 g, 0.986 mmol) and H-
Lys(Cbz)-OtBU HCl (0.242 g, 0.655 mmol) were dis-
solved in DMF (6.0 ml). This solution was cooled to
0 ꢁC, then a DMF solution of PyBrop (0.362 g,
0.777 mmol) and a DIEA (0.57 ml, 3.26 mmol) was add-
ed and stirred for 2 h at 0 ꢁC and another 21 h at room
temperature. The reaction mixture was poured into
water (200 ml) and stored in a refrigerator overnight.
Resultant precipitate was collected by centrifugation
and dried in vacuum. The product was obtained as a
white powder, 0.196 g, 0.390 mmol, 59.7%. Rf = 0.83
4.31. 60-N-[aN[(Guanine-9-yl)acetyl]-NG-(2,2,4,6,7-pen-
tamethyl-dihydrobenzofuran-5-sulfonyl) arginyl]neamine
(Gb-R(Pbf)-neamine, 27)
1
Compound 26 (0.050 g, 0.081 mmol) was dissolved in
DMF (1.5 ml) and cooled to 0 ꢁC, WSC HCl
(0.018 g, 0.097 mmol). Neamine (0.078 g, 0.243 mmol)
was dissolved in water (1.5 ml), the reaction mixture
of compound 13 added slowly and stirred overnight.
The reaction mixture was concentrated and dried.
The compound 14 (mixture of D and L forms) was
obtained as a white powder. This crude product
was dissolved in 10% acetic acid solution and puri-
fied by HPLC. Gb-(D)R(Pbf)-Neamine. Yield:
10.6 mg, 13.1 lmol, 16%. Rf = 0.275 (solvent 4), 1H
NMR (D2O, 300 MHz) d 7.92 (s, 1H), d 4.90 (d,
1H), d 4.77 (s, 2H), d 3.85 (t, 1H), d 3.65–2.96
(m, majority), d 2.86 (sm, 2H), d 2.35 (s, 3H), d
2.28 (s, 3H), d 1.89 (s, 3H), d 1.28 (s, 6H). Gb-
(L)R(Pbf)-Neamine. Yield: 12.1 mg, 15.0 lmol, 19%.
(solvent 2), H NMR (DMSO-d6, 300 MHz) d 11.25 (s
1H), d 8.48 (d, 1H), d 7.42 (s, 1H), d 7.35 (m, 6H), d
5.00 (s 2H), d 4.33 (s, 2H), d 4.09 (m, 1H), d 2.98 (m,
2H), d 1.74 (s 3H), d 1.62 (m, 2H), d 1.39 (s, 9H), d
1.73 (s, 3H), d 1.31 (m, 4H).
a
4.34. N[(Thymine-1-yl)acetyl]-eN-(benzyloxycarbonyl)lysi-
nate (Tb-K(Cbz)-OH, 30)
Compound 29 (0.090 g, 0.179 mmol) was dissolved in
dichloromethane (1.5 ml) and TFA (1.5 ml) was added
and stirred for 30 min. at 0 ꢁC and 3 h at room temper-
ature. The reaction mixture was concentrated. The crude
product was crystallized from cold ethanol. White pre-
cipitate was collected and dried. Yield 0.079 g,
0.178 mmol, 99.2%. Rf = 0.54 (solvent 4).