M. Gao et al. / Bioorg. Med. Chem. Lett. 26 (2016) 1371–1375
1375
(
d) 2-Chloro-N-(4-hydroxy-3-methoxyphenyl)acetamide (1c): Compound 1c was
d 4.33 (br s, 2H, NH
([M+H] , 100%).
2
), 6.45 (br s, 4H, 2 Â NH
2
), 7.68 (s, 1H, Ar-H). MS (ESI): 126
+
prepared from 4-amino-2-methoxyphenol and 2-chloroacetyl chloride accord-
ing to the procedure for the synthesis of 1b, as a brown solid, yield 62%.
(j) 6-Amino-7,9-dihydro-8H-purine-8-thione (7): To a mixture of compound 6
(4.13 g, 33 mmol) in solvents of EtOH (200 mL) and water (10 mL) was added
KOH (2.28 g, 35 mmol) and CS (3.33 g, 43.8 mmol). The reaction mixture was
2
stirred at reflux for overnight. The solvents were evaporated, and the residue
1
R
3
8
f
= 0.65 (1:19 MeOH/CH
H, OCH ), 4.19 (s, 2H, CH
.5 Hz, 1H, Ph-H), 7.24 (d, J = 2.0 Hz, 1H, Ph-H), 8.79 (s, 1H, NH), 10.06 (s, 1H,
2
Cl
2
2
), mp 126–128 °C. H NMR (DMSO-d
), 6.70 (d, J = 8.5 Hz, 1H, Ph-H), 6.94 (dd, J = 2.5,
6
): d 3.71 (s,
3
+
À
OH). MS (ESI): 216 ([M+H] , 70%); MS (ESI): 214 ([MÀH] , 100%).
e) 6-Methoxypyridine-2,3-diamine (2): To a solution of 6-methoxy-3-nitropy-
ridin-2-amine (25 g, 0.148 mol) in methanol was added palladium on carbon
10% Pd/C, 0.70 g). The reaction mixture was stirred at rt for 15 h under an
were rinsed with water and acetone to obtain a yellow product 7 (3.64 g, 66%).
1
(
R
NH
f
= 0.24 (1:9 MeOH/CH
2
Cl
2
), mp >350 °C. H NMR (DMSO-d
6
): d 6.59 (s, 2H,
+
2
), 7.98 (s, 1H, Ar-H), 12.20 (br s, 2H, 2 Â NH). MS (ESI): 168 ([M+H] , 90%);
À
(
MS (ESI): 166 ([MÀH] , 100%).
atmosphere of hydrogen gas (55 psi). The mixture was filtered over Celite, and
the filtrate was collected. The solvent was evaporated to afford a black solid
crude product 2 (20.4 g). Because of the instability of the diamino analog, the
(k) General procedure to prepare compounds 8a–c: To a solution of compound 7
(334 mg, 2.0 mmol) in a mixture of EtOH (100 mL) and 1 N aq NaOH (2.6 mL,
2.6 mmol), compound 1 (2.0 mmol) was added. The resulting reaction mixture
was stirred overnight at 60 °C. Then solvents were removed under reduced
pressure, and the crude product was purified by column chromatography on
product was immediately used in next step reaction without further purifi-
+
cation. R
f
= 0.40 (1:19 MeOH/CH
2
Cl
2
). MS (ESI): 140 ([M+H] , 100%).
(
f) 5-Methoxy-1,3-dihydro-2H-imidazo[4,5-b]pyridine-2-thione (3): To a solution
of compound 2 (20.56 g, 148 mmol) in a mixture of EtOH (200 mL) and water
40 mL) was added KOH (9.75 g, 148 mmol) and CS (22.5 g, 296 mmol). The
2 2
silica gel with eluent (2:98 to 10:90 MeOH/CH Cl ) to give a gray solid product
8 (45-60% yields).
(
2
2-((6-Amino-9H-purin-8-yl)thio)-N-(3,4-dimethoxyphenyl)acetamide (8a):
1
reaction mixture was stirred at reflux for 12 h. The solvents were evaporated,
and the residue was purified by column chromatography on silica gel with
eluent (2:98 to 10:90 MeOH/CH
6
3
1
(
(
2
R
f
= 0.26 (1:9 MeOH/CH
6H, 2 Â OCH ), 4.12 (s, 2H, CH
Ar-H and NH ), 7.25 (d, J = 2.5 Hz, 1H, Ar-H), 8.05 (s, 1H, Ar-H), 10.18 (s, 1H,
2
Cl
2
), mp 270–272 °C. H NMR (DMSO-d
2
6
): d 3.70 (s,
3
), 6.87 (d, J = 8.5 Hz, 1H, Ar-H), 7.08–7.10 (m, 3H,
2
Cl
Cl
), 6.56 (d, J = 8.5 Hz, 1H, Ar-H), 7.44 (d, J = 8.5 Hz, 1H, Ar-H),
2
), affording a gray solid product 3 (16.9 g,
2
1
+
À
3%). R
.83 (s, 3H, OCH
f
= 0.50 (1:19 MeOH/CH
2
2
), mp 246–248 °C. H NMR (DMSO-d
6
): d
NH), 13.14 (s, 1H, NH). MS (ESI): 361([M+H] , 100%); MS (ESI): 359 ([MÀH] ,
3
30%).
+
À
2.73 (s, 2H, NH). MS (ESI): 182 ([M+H] , 100%); MS (ESI): 180 ([MÀH] , 23%).
2-((6-Amino-9H-purin-8-yl)thio)-N-(3-hydroxy-4-methoxyphenyl)acetamide
1
g) General procedure to prepare compounds 4a–c: To a solution of compound 3
(8b): R
(s, 3H, OCH
8.5 Hz, 1H, Ar-H), 7.05 (br s, 2H, NH
Ar-H), 9.05 (s, 1H, OH), 10.57 (s, 1H, NH), 13.09 (s, 1H, NH). MS (ESI): 347([M
f
= 0.29 (1:7 MeOH/CH
2
Cl
2
2
), mp 261–263 °C. H NMR (DMSO-d
6
): d 3.70
360 mg, 2.0 mmol) in a mixture of EtOH (60 mL) and 1 N aq NaOH (2.2 mL,
.2 mmol), compound 1 (2.1 mmol) was added. The resulting reaction mixture
3
), 4.12 (s, 2H, CH
), 6.81 (d, J = 8.5 Hz, 1H, Ar-H), 6.90 (dd, J = 2.5,
) 7.14 (d, J = 2.5 Hz, 1H, Ar-H), 8.05 (s, 1H,
2
was stirred overnight at rt. Then solvents were removed under reduced
pressure, and the crude product was purified by column chromatography on
silica gel with eluent (0:100 to 3:97 MeOH/CH
+
À
+H] , 100%); MS (ESI): 345 ([MÀH] , 70%). HRMS (ESI): calcd for C14
15 6 3
H N O S
+
2
Cl
2
) to give a white solid
([M+H] ) 347.0926, found 347.0912.
product 4 (50–65% yields).
N-(3,4-Dimethoxyphenyl)-2-((5-methoxy-3H-imidazo[4,5-b]pyridin-2-yl)thio)ac-
2-((6-Amino-9H-purin-8-yl)thio)-N-(4-hydroxy-3-methoxyphenyl)acetamide
1
(8c): R
(s, 6H, 2 Â OCH
J = 2.0, 8.5 Hz, 1H, Ar-H), 7.09 (br s, 2H, NH
(s, 1H, Ar-H), 8.77 (s, 1H, OH), 10.09 (s, 1H, NH), 13.11 (s, 1H, NH). C NMR
(DMSO-d ): d 35.89, 55.48, 103.50, 104.72, 106.40, 108.10, 111.79, 115.19,
f
= 0.37 (1:7 MeOH/CH
2
Cl
2
), mp 266–268 °C. H NMR (DMSO-d
), 6.65 (d, J = 8.5 Hz, 1H, Ar-H), 6.93 (dd,
), 7.23 (d, J = 2.0 Hz, 1H, Ar-H), 8.05
6
): d 3.71
1
etamide (4a): R
): d 3.66 (s, 6H, 2 Â OCH
J = 8.5 Hz, 1H, Ar-H), 6.88 (d, J = 8.5 Hz, 1H, Ar-H), 7.07 (d, J = 8.0 Hz, 1H, Ar-H),
f
= 0.61 (1:19 MeOH/CH
2
Cl
2
), mp 175–177 °C. H NMR (DMSO-
3
), 4.11 (s, 2H, CH
2
d
6
3
), 3.84 (s, 3H, OCH
3
), 4.20 (s, 2H, CH ), 6.59 (d,
2
2
13
7
.28 (d, J = 2.5 Hz, 1H, Ar-H), 7.80 (d, J = 8.5 Hz, 1H, Ar-H), 10.30 (br s, 1H, NH),
6
+
+
1
2.76 and 13.10 (2s, 1H, NH). MS (ESI): 375 ([M+H] , 100%); MS (ESI): 373
119.20, 130.89, 142.73, 142.90, 147.23, 151.57, 165.44. MS (ESI): 347 ([M+H] ,
À
À
(
[MÀH] , 20%).
80%); MS (ESI): 345 ([MÀH] , 100%). HRMS (ESI): calcd for C14
15 6 3
H N O S ([M
+
N-(3-Hydroxy-4-methoxyphenyl)-2-((5-methoxy-3H-imidazo[4,5-b]pyridin-2-yl)
+H] ) 347.0926, found 347.0914.
1
11
thio)acetamide (4b): R
f
= 0.35 (1:19 MeOH/CH
2
Cl
2
), mp 216–218 °C. H NMR
), 4.19 (s, 2H, CH ), 6.59 (d,
(l) General procedure to prepare target tracers N-(3-[ C]methoxy-4-methoxyphe-
11
(
DMSO-d ): d 3.71(s, 3H, OCH
6
3
), 3.85 (s, 3H, OCH
3
2
nyl)-2-((5-methoxy-3H-imidazo[4,5-b]pyridin-2-yl)thio)acetamide (3-[ C]4a)
11
J = 8.5 Hz, 1H, Ar-H), 6.82 (d, J = 8.5 Hz, 1H, Ar-H), 6.92 (dd, J = 2.0, 8.5 Hz, 1H,
Ar-H), 7.14 (d, J = 2.5 Hz, 1H, Ar-H), 7.73 and 7.80 (2s, 1H, Ar-H), 9.06 (s, 1H,
OH), 10.17 (br s, 1H, NH), 12.74 and 13.09 (2s, 1H, NH). C NMR (DMSO-d
and N-(4-[ C]methoxy-3-methoxyphenyl)-2-((5-methoxy-3H-imidazo[4,5-b]pyr-
11 11
idin-2-yl)thio)acetamide (4-[ C]4a); 2-((6-amino-9H-purin-8-yl)thio)-N-(3-[ C]
1
3
11
6
): d
methoxy-4-methoxyphenyl)acetamide (3-[ C]8a) and 2-((6-amino-9H-purin-8-
yl)thio)-N-(4-[ C]methoxy-3-methoxyphenyl)acetamide (4-[ C]8a):
was produced by the N(p,
(9.5 cm ) aluminum gas target provided with the Siemens RDS-111 Eclipse
cyclotron. The target gas consisted of 1% oxygen in nitrogen purchased as a
specialty gas from Praxair, Indianapolis, IN. Typical irradiations used for the
1
1
11
11
3
1
5.99, 53.34, 55.88, 104.56, 107.62, 109.83, 112.54, 121.00, 128.20, 130.10,
[
C]CO
2
+
14
11
32.50, 143.97, 146.48, 148.00, 159.96, 165.37. MS (ESI): 361([M+H] , 100%);
a)
C nuclear reaction in the small volume
À
+
3
MS (ESI): 359 ([MÀH] , 25%). HRMS (ESI): calcd for C16
61.0971, found 361.0954.
N-(4-Hydroxy-3-methoxyphenyl)-2-((5-methoxy-3H-imidazo[4,5-b]pyridin-2-yl)
H
17
N
4
O
4
S ([M+H] )
3
1
thio)acetamide (4c): R
DMSO-d ): d 3.72(s, 3H, OCH
), 6.60 (t, J = 8.5 Hz, 1H, Ar-H), 6.68 (d, J = 8.5 Hz, 1H, Ar-H), 6.91 and
.94 (2d, J = 8.5, 1H, Ar-H), 7.26 (s, 1H, Ar-H), 7.71 and 7.81 (2d, J = 8.5 Hz, 1H,
f
= 0.33 (1:19 MeOH/CH
2
Cl
2
), mp 119–121 °C. H NMR
development were 50
run produced approximately 25.9 GBq of
hydroxyl precursor 4b (4c, 8b or 8c) (0.1–0.3 mg) was dissolved in CH
(300 L). To this solution was added 2 N NaOH (2 L). The mixture was
transferred to a 5-mL small reaction vial. No-carrier-added (high specific
lA beam current for 15 min on target. The production
11
(
6
3
), 3.84 and 3.85 (2s, 3H, OCH
3
), 4.18 and 4.21 (2s,
[
C]CO
2
at EOB. The phenolic
CN
2
6
H, CH
2
3
l
l
Ar-H), 8.77 (s, 1H, OH), 10.18 and 10.24 (2br s, 1H, NH), 12.74 and 13.10 (2s,
H, NH). 13C NMR (DMSO-d
21.30, 122.05, 128.04, 130.22, 131.00, 142.70, 147.24, 147.92, 159.94, 165.29.
): d 35.91, 53.37, 55.47, 104.04, 104.69, 115.24,
activity) [ C]CH OTf (13.9 GBq) that was produced by the gas-phase produc-
11
1
1
6
3
tion method11 within 11 min from
(21.8 GBq) and [ C]CH Br (13.9 GBq) with silver triflate (AgOTf) column was
3
[
1
1
1
1
2 4
C]CO (25.9 GBq) through [ C]CH
+
À
11
MS (ESI): 361([M+H] , 100%); MS (ESI): 359 ([MÀH] , 30%). HRMS (ESI): calcd
+
for C16
H
N
17 4
O
4
S ([M+H] ) 361.0971, found 361.0955.
h) 5-Nitrosopyrimidine-4,6-diamine (5): A solution of 4,6-pyrimidinediamine
16.0 g, 110 mmol) in 3 N HCl (367 mL) was cooled in an ice bath, and then
(8.73 g, 126 mmol) in H O (30 mL) was added dropwise. The mixture
was stirred for 30 min at 0 °C, and then 2 h at rt. The violet solution was
neutralized with NaHCO to pH 8, the resulting precipitate collected,
suspended in H O and head to 70 °C, filtered, washed with MeOH, and dried
to give a blue solid product 5 (14.5 g, 78%). R = 0.81 (1:5 MeOH/CH Cl ), mp
): d 7.93 (s, 1H, Ar-H), 8.04 (s, 1H, NHH), 8.44 (s, 1H,
passed into the reaction vial at rt until radioactivity reached a maximum
(2 min), and then the reaction vial was isolated and heated at 80 °C for 3 min.
The contents of the reaction vial were diluted with NaHCO (0.1 M, 1 mL). The
3
reaction vial was connected to a C-18 Plus Sep-Pak cartridge. The labeled
product mixture solution was passed onto the cartridge for SPE purification by
(
(
NaNO
2
2
3
gas pressure. The cartridge was washed with H
2
O (2 Â 3 mL), and the aqueous
2
washing was discarded. The product was eluted from the cartridge with EtOH
(2 Â 2 mL), and then passed onto a rotatory evaporator. The solvent was
removed by evaporation (3 min) under vacuum. The final volume of ethanol
after evaporation was ꢀ1 mL. The labeled product was reformulated with
f
2
2
1
>
350 °C. H NMR (DMSO-d
NHH), 9.11 (s, NHH), 10.05 (s, 1H, NHH). MS (ESI): 140 ([M+H] , 100%).
i) Pyrimidine-4,5,6-triamine (6): Compound 5 (10.0 g, 72 mmol) was dissolved
6
+
(
saline (10 mL), sterile-filtered through a sterile vented Millex-GS 0.22 lm
in water (110 mL) and treated with sodium dithionite (27.6 g, 158 mmol),
which was added in portions at rt. The resulting yellow mixture was then
cellulose acetate membrane and collected into a sterile vial. Total radioactivity
(4.7–7.1 GBq) was assayed and the total volume (10–11 mL) was noted for
tracer dose dispensing. The overall synthesis time including SPE purification
and reformulation was 23 min. The radiochemical yields decay corrected to
treated with 50% aq H
and then cooled to 10 °C in an ice bath. The resulting precipitate was filtered
off and washed with aq. ethanol and dried to afford yellowish solid
2 4
SO (141 g, 720 mmol) and heated to 80 °C for 5 min,
a
EOB, from [11C]CO
, were 50–60%. The same procedure was used to prepare the
2
11 11 11 11
intermediate 4,5,6-triaminopyrimidine sulfate (8.81 g). A mixture of 4,5,6-
triaminopyrimidine sulfate (8.81 g, 39.5 mmol), NaOH (3.48 g, 87 mmol) and
water (80 mL) was heated to 80 °C until all the solid was dissolved. The
solution was cooled to 0 °C and pH was adjusted to 7.0 with 1 N HCl,
target tracers 3-[ C]4a and 4-[ C]4a, 3-[ C]8a and 4-[ C]8a from their
corresponding precursors 4b and 4c, 8b and 8c. Retention times in the
analytical HPLC system were:
t
R
4b = 5.42 min,
t
R
4c = 5.35 min,
t
t
R
R
R
11
11
4a = 7.74 min,
t
R
3-[ C]4a = 7.85 min,
t
R
4-[ C]4a = 7.79 min; and
11
whereupon the free base precipitated as an off-white solid product 6 (3.69 g,
8b = 3.57 min, t
4-[ C]8a = 5.61 min.
R
8c = 3.46 min, t 8a = 5.53 min, t
R
R
3-[ C]8a = 5.68 min, t
1
11
4
1% from 5). R
f
= 0.26 (1:5 MeOH/CH
2
Cl
2
), mp 288–290 °C. H NMR (DMSO-d
6
):