2,4-Diaminopyrimidine Derivatives: DHFR Inhibitors
J . Org. Chem., Vol. 64, No. 2, 1999 445
13C NMR (CDCl3, 75.6 MHz) δ 14.7, 18.9, 21.5, 27.6, 30.6, 30.8,
35.4, 40.4, 40.5, 62.1, 108.4, 155.3, 156.0, 163.3, 176.1; MS m/e
(relative intensity) 370 (11), 368 (28), 327 (16), 325 (43), 309
(54), 307 (100); HRMS calcd for C17H25N4OSCl 368.1437, found
368.1435. Anal. Calcd for C17H25N4OSCl: C, 55.35; H, 6.83;
N, 15.19. Found: C, 55.41; H, 6.56; N, 14.88.
2,4-Dia m in o-5,6,7,8,9,10-h exa h yd r o-5,9-m eth a n op yr im -
id o[4,5-b]a zocin e (3). Meth od A. A suspension of 15 (54 mg,
0.19 mmol) and LiOH (54 mg, 0.12 mmol, 7 equiv) in 5 mL of
MeOH and 0.5 mL of water was heated to 50 °C. The resulting
pale yellow solution was held at this temperature for 2.5 h,
cooled to rt, and concentrated in vacuo. The resulting solid
residue was purified by reversed-phase HPLC (C-18 prepara-
tive column) using a gradient of 1% MeCN/0.1% TFA in water
and 90% MeCN/0.1% TFA in water. The resulting hygroscopic
white solid was suspended in water and dissolved by slowly
adjusting the pH to 9 with 0.1 M K2CO3 in water. The resulting
solution (10 mL) was extracted with CHCl3 (4 × 5 mL), and
the combined organic extracts were dried by filtration through
a pad of Na2SO4. The filtrate was concentrated in vacuo to
afford a white microcrystalline solid (20 mg, 50%): mp 216-
218 °C; 1H NMR (CDCl3, 500 MHz) δ 1.28 (m, 1 H), 1.37 (br d,
1 H, J ) 13.1 Hz), 1.45 (m, 3 H), 1.61 (m, 2 H), 1.73 (d, 1 H,
J ) 11.9 Hz), 2.84 (br s, 1 H), 3.48 (br s, 1 H), 5.10 (br s, 2 H),
5.38 (br s, 2 H), 6.35 (d, 1 H, J ) 2.9 Hz); 13C NMR (CDCl3,
75.6 MHz) δ 17.5, 25.2, 30.2, 30.9, 34.5, 44.7, 84.9, 159.6, 160.8,
161.4; MS m/e (relative intensity) 205 (24), 162 (100), 145 (11);
HRMS calcd for C10H15N5 205.1327, found 205.1331. Anal.
Calcd for C10H15N5: C, 58.52; H, 7.37; N, 34.12. Found: C,
58.53; H, 7.22; N, 34.02.
2-P ivaloylam in o-4-azido-9-eth ylth io-5,6,7,8,9,10-h exah y-
d r o-5,9-m eth a n op yr im id o[4,5-b]a zocin e (13). A magneti-
cally stirred suspension of 12 (280 mg, 0.76 mmol) and TMSN3
(174 mg, 1.50 mmol, 2.0 equiv) in 5 mL of N-methylpyrroli-
dinone was heated at 110 °C for 6 h. After the reaction mixture
was allowed to cool to rt, the solvent was removed by
distillation under reduced pressure using a short-path ap-
paratus. The resulting semisolid residue was purified by radial
chromatography (2-mm plate) using a gradient of 2-5% MeOH
in CH2Cl2 as eluent to afford 256 mg (90%) of a white foam:
1
mp 45-60 °C; H NMR (CDCl3, 500 MHz) δ 1.21 (t, 3 H, J )
7.4 Hz), 1.31 (s, 9 H), 1.41 (m, 1 H), 1.56-2.04 (m, 7 H), 2.65
(m, 2 H), 3.14 (m, 1 H), 5.65 (s, 1 H), 7.71 (s, 1 H); 13C NMR
(CDCl3, 75.6 MHz) δ 14.6, 18.8, 21.4, 27.5, 28.2, 30.6, 35.3,
40.2, 40.5, 61.9, 108.3, 155.2, 156.6, 176.0; IR (NaCl) 3245,
2131, 1700, 1604, 1575 cm-1; MS m/e (relative intensity) 375
(22), 332 (16), 314 (53), 288 (31); HRMS calcd for C17H25N7OS
375.1841, found 375.1829.
2,4-Diam in o-9-h ydr oxy-5,6,7,8,9,10-h exah ydr o-5,9-m eth -
a n op yr im id o[4,5-b]a zocin e (16). A suspension of 2,4,6-
triaminopyrimidine (0.31 g, 2.5 mmol) and 2-cyclohexen-1-one
in glacial acetic acid (10 mL) was placed in a sealed tube which,
upon heating at 120 °C for 20 min, gave a clear solution. The
reaction mixture was heated for another 17 h at 120 °C. The
solvent was removed in vacuo to give a white solid that was
purified by column chromatography on silica gel using 50%
MeOH/CH2Cl2 as eluent to give 0.23 g (41%) of 16: mp 230
°C; 1H NMR (DMSO-d6, 500 MHz) δ 1.30 (m, 2 H), 1.46 (m, 1
H), 1.49 (m, 2 H), 1.56 (m, 1 H), 1.63 (d, 1 H, J ) 11.6 Hz),
1.69 (d, 1 H, J ) 11.6 Hz), 2.93 (s, 1 H), 5.18 (s, 2 H), 5.43 (s,
2 H), 5.52 (s, 1 H), 5.93 (s, 1 H); 13C NMR (DMSO-d6, 125 MHz)
δ 19.9, 28.2, 30.4, 41.2, 79.6, 84.7, 159.8, 161.1, 161.2; IR (KBr)
3371, 3189, 2971, 2858, 1611 cm-1; MS m/e (relative intensity)
221 (36), 178 (100); HRMS calcd for C10H15N5O 221.1276, found
221.1284.
2,4-Bis(pivaloylam in o)-9-(pivaloyloxy)-5,6,7,8,9,10-h exa-
h yd r o-5,9-m eth a n op yr im id o[4,5-b]a zocin e (19). A suspen-
sion of 16 (1.0 g, 0.46 mmol) in 4 mL of pivalic anhydride was
heated at 140 °C for 6 h. The solvent was removed in vacuo
with the aid of a short-path distillation apparatus. The
resulting solid residue was purified by column chromatography
using 50% EtOAc in hexane as eluent to give 1.84 g (85%) of
19: mp 145-147 °C; 1H NMR (DMSO-d6, 500 MHz) δ 1.15 (s,
9 H), 1.17 (s, 9 H), 1.18 (s, 9 H), 1.54 (m, 2 H), 1.60 (m, 2 H),
1.93 (m, 1 H), 2.05 (m, 1 H), 2.10 (d, 1 H, J ) 11.3 Hz), 2.16
(d, 1 H, J ) 11.3 Hz), 3.09 (s, 1 H), 7.92 (s, 1 H), 9.31 (s, 1 H),
9.50 (s, 1 H); 13C NMR (DMSO-d6, 125 MHz) δ 19.1, 26.7, 26.8,
27.1, 28.9, 30.0, 32.5, 37.1, 38.7, 40.0, 87.7, 107.6, 154.2, 155.9,
162.4, 174.8, 175.7, 176.5, 176.7; IR (KBr) 3414, 3273, 2956,
1710, 1590 cm-1; MS m/e (relative intensity) 473 (50), 372 (36),
314 (100), 286 (59); HRMS calcd for C25H39N5O4 473.3002,
found 473.3017. Anal. Calcd for C25H39N5O4: C, 63.40; H, 8.30;
N, 14.79. Found: C, 63.56; H, 8.34; N, 14.26.
2,4-Bis(pivaloylam in o)-5,6,7,8,9,10-h exah ydr o-5,9-m eth -
a n op yr im id o[4,5-b]a zocin e (20). To a solution of 19 (0.95
g, 2.0 mmol) in 10 mL of pyridine was added sodium cy-
anoborohydride (0.50 g, 8.0 mmol) in one portion, and the
reaction mixture was heated at 110 °C for 24 h. The solvent
was removed under reduced pressure followed by the addition
of water. The aqueous layer was extracted with CH2Cl2 (2 ×
20 mL) and dried over Na2SO4. The solvent was removed in
vacuo, and the residue obtained was purified by column
chromatography using 5% MeOH in CH2Cl2 as eluent to give
19 (0.66 g, 88%) as a white powder: mp 160-161 °C; 1H NMR
(DMSO-d6, 500 MHz) δ 1.17 (s, 9 H), 1.18 (s, 9 H), 1.42 (m, 3
H), 1.50 (m, 2 H), 1.67 (m, 2 H), 1.80 (d, 1 H, J ) 12.5 Hz),
2.91 (s, 1 H), 3.61 (s, 1 H), 7.64 (d, 1 H, J ) 4.2 Hz), 9.18 (s, 1
H), 9.33 (s, 1 H); 13C NMR (DMSO-d6, 125 MHz) δ 17.0, 26.6,
26.9, 27.1, 28.3, 31.1, 33.4, 38.6, 40.0, 44.8, 107.9, 153.5. 155.3.
2-P iva loyla m in o-4-a m in o-9-eth ylth io-5,6,7,8,9,10-h exa -
h yd r o-5,9-m eth a n op yr im id o[4,5-b]a zocin e (14). A mag-
netically stirred suspension of 13 (256 mg, 0.68 mmol) and
one weight equivalent of 5% Pd/C in 5 mL of EtOH was
reduced under one atmosphere of H2 for 3 h. The reaction
mixture was filtered through Celite and the filter cake was
washed with 15 mL of hot EtOH. Concentration of the filtrate
in vacuo gave an oil which was purified by radial chromatog-
raphy (1-mm plate) using 10% MeOH and 1% Et3N in CH2Cl2
as eluent. The resulting clear, colorless oil (214 mg, 90%)
1
solidified upon standing: mp 130 °C (dec); H NMR (CDCl3,
500 MHz) δ 1.20 (t, 3 H, J ) 7.4 Hz), 1.26 (s, 9 H), 1.27 (m, 1
H, obscured), 1.49 (m, 1 H), 1.56 (m, 3 H), 1.75 (m, 2 H), 1.85
(d, 1 H, J ) 12.2 Hz), 2.62 (m, 2 H), 2.89 (m, 1 H), 4.68 (s, 2
H), 5.33 (s, 1 H), 7.63 (s, 1 H); 13C NMR (CDCl3, 75.6 MHz) δ
14.7, 19.1, 21.3, 27.5, 28.2, 29.9, 36.1, 40.1, 41.1, 61.6, 90.2,
155.3, 159.4, 161.7, 175.9; MS m/e (relative intensity) 349 (19),
306 (19), 288 (100), 57 (40); HRMS calcd for C17H27N5OS
349.1936, found 349.1931.
2-P iva loyla m in o-4-a m in o-5,6,7,8,9,10-h exa h yd r o-5,9-
m eth a n op yr im id o[4,5-b]a zocin e (15). A 50-mL, single-
necked, pear-shaped flask fitted with a stoppered Claisen head,
reflux condenser, and gas inlet and containing a solution of
14 (150 mg, 0.43 mmol), Bu3SnH (150 mg, 0.52 mmol, 1.2
equiv), and AIBN (35 mg, 0.22 mmol, 0.5 equiv) in 10 mL of
toluene, freshly distilled from CaH2, was immersed in an oil
bath heated to 125 °C. The flask was carefully evacuated and
purged with Ar, a process that was repeated three times.
Previous experiments indicated that periodic reinitiation of the
reaction was required. At 6-h intervals, over a 36-h period,
the reaction vessel was removed from the oil bath and
additional AIBN (35 mg, 0.22 mmol, 0.5 equiv) was added to
the reaction mixture. The evacuation and purging routine was
repeated, and the reaction was allowed to continue until TLC
analysis (1% Et3N and 5% MeOH in CH2Cl2) indicated that
the starting material had been consumed. Removal of the
solvent in vacuo followed by Kugelrohr distillation of the oily
residue afforded a semisolid material which was purified by
radial chromatography using 1% Et3N and 5% MeOH in
CH2Cl2 as eluent. The desulfurized material was obtained as
a white solid (100 mg, 80%): mp 220-221 °C; 1H NMR (CDCl3,
500 MHz) δ 1.23 (m, 1 H, partially obscured), 1.27 (s, 9 H),
1.40 (dt, 1 H, J ) 4.3, 12.9 Hz), 1.49 (d, 1 H, J ) 11.7 Hz),
1.59 (m, 2 H), 1.72 (d, 2 H, J ) 11.6 Hz), 1.84 (d, 1 H, J ) 12.2
Hz), 2.79 (br s, 1 H), 3.69 (br s, 1 H), 4.58 (br s, 2 H), 5.46 (br
s, 1 H), 7.62 (br s, 1 H); 13C NMR (CDCl3, 75.6 MHz) δ 17.8,
26.6, 27.7, 29.9, 30.7, 34.7, 40.2, 46.3, 91.2, 154.9, 159.1, 161.6,
176.1; MS m/e (relative intensity) 289 (10), 246 (13), 205 (27),
189 (9), 162 (41); HRMS calcd for C15H23N5O 289.1902, found
289.1916.