6686 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 21
Wu et al.
interactions with MBP. The binding mechanism is not known
but our previous work with structurally related myelin-imaging
probes indicated that they bind to pleated ꢀ-sheet structure
present in myelin basic protein (MBP).18
1.5. Synthesis of N-Methyl-{4-[2-(4-amino-phenyl)-vinyl]-phen-
yl}-amine (7). To a solution of 4 (50 mg, 2 mmol) dissolved in
THF (5 mL) under argon was added to Et3N (1 mL). The solution
was stirred for 4 h. The solvent was evaporated under vacuum,
and the protected product was then used without further purification.
To the solution of the above protected product dissolved in DMF
(5 mL) were added NaH (0.10 g) and iodomethane (1 mL). The
vial was sealed and stirred overnight. Then the solution was diluted
with methanol (8 mL) and 1 M NaOH solution (2 mL). After stirred
for another 2 h, the solution was extracted with ethyl acetate. The
combined organic layer was washed with water and brine and dried
over Na2SO4. Following concentration, the reduced product was
then subsequently used without further purification.
Conclusion
Several stilbene derivatives have been synthesized and
evaluated for potential in vivo imaging of myelinated brain
regions. These small molecular probes selectively stain myeli-
nated white matter regions such as the corpus callosum and
cerebellum in mouse brain tissue sections. The white matter
staining was also achieved in situ following tail vein injections,
suggesting they readily enter the brain. Both spectrophotometry
and radioligand-based binding assays suggested a specific
interaction of these compounds with myelin membranes. These
in vitro and in situ binding properties, coupled with its
permeability across the BBB, make them promising candidates
for further investigation as in vivo myelin-imaging agents.
To the suspension of the above compound in acetic acid (10
mL) was added tin(II) chloride (1.0 g). The suspension was heated
to reflux for 2 h. After concentration, the residue was dissolved in
ethyl acetate, washed with 2N NaOH solution, water, and brine.
Dried over Na2SO4, the solution was concentrated and purified by
flash column (hexanes:ethyl acetate ) 2:1 to 1:1) to give 15 mg of
1
7 (0.6 mmol, 35% yield for the above three steps). H NMR (400
MHz, CDCl3): 7.36 (d, J ) 8.3 Hz, 2 H), 7.32 (d, J ) 8.2 Hz, 2
H), 6.87 (AB, J ) 18.7 Hz, 16.5 Hz, 2 H), 6.69 (d, J ) 8.2 Hz, 2
H), 6.62 (d, J ) 8.3 Hz, 2 H), 3.92 (br, 3 H), 2.88 (s, 2 H). HR-
ESIMS: m/z calcd for C15H16N2 (M + H+), 225.1386; found,
225.1385. Melting point: 143.7-144.7 °C.
1.6. Synthesis of 4-Amino-4′-iodostilbene (8). To a solution
of diethyl 4-nitrobenzylphosphate (0.44 g, 1.61 mmol) dissolved
in DMF (10 mL) was added NaH (0.07 g, 1.75 mmol). The
suspension was stirred for 1 h followed by addition of 4-iodo-
benzaldehyde (0.35 g, 1.51 mmol). The suspension was stirred for
another 2 h. Water was added and the solid was collected by
filtration to give 8 (0.40 g, 1.14 mmol, yield: 75%). 1H NMR (400
MHz, CDCl3): 8.24 (d, J ) 8.65 Hz, 2 H), 7.88 (d, J ) 8.68 Hz,
2 H), 7.80 (d, J ) 8.19 Hz, 2 H), 7.49 (m, J ) 8.2 Hz, 4 H).
1.7. Synthesis of 4-[2-(4-Iodo-phenyl)-vinyl]-phenylamine (9).
To a suspension of compound 8 (0.20 g, 0.57 mmol) in ethanol
(10 mL) was added Tin(II) chloride (1.00 g, 5 mmol) and heated
to reflux for 4 h under argon. The ethanol was evaporated under
vacuum. The residue was dissolved in ethyl acetate, washed with
1N NaOH, water, and brine. Dried over Na2SO4, the solution was
concentrated and purified by flash column (hexanes:ethyl acetate
) 2:1 to 1:1) to give 9 (0.18 g, quant. yield). 1H NMR (400 MHz,
CDCl3): 7.66 (d, J ) 8.25 Hz, 2 H), 7.35 (d, J ) 8.34 Hz, 2 H),
7.04 (d, J ) 15.73 Hz, 1 H), 6.71 (d, J ) 16.26 Hz, 1 H), 6.70 (d,
J ) 8.29 Hz, 2 H). HR-ESIMS: m/z calcd for C14H12IN (M + H+),
322.0087; found, 322.0084. Melting point: 213.4-215.2 °C.
Methods and Materials
1. Synthesis. 1.1. Synthesis of 4,4′-Dinitro-trans-stilbene (2).
Under Ar, (4-nitro-benzyl)-phosphoric acid diethyl ester (1, 1.81 g,
6.6 mmol) and 4-nitrobenzaldehyde (1.00 g, 6.6 mmol) were
dissolved in DMF (10 mL) and EtOH (10 mL). Then NaOCH3
(2.3 mL, 4.37 M) in MeOH was added and the suspension was
stirred for another 3 h. The solid was filtered and dried in vacuum
to give 1.50 g (yield: 83%) of 4,4′-dinitro-trans-stilbene. 1H NMR
(300 MHz, CDCl3): 8.28 (d, J ) 8.4 Hz, 4 H), 7.94 (d, J ) 8.4 Hz,
4 H), 7.69 (s, 2 H).
1.2. Synthesis of 4,4′-Diamino-trans-stilbene (3) and 4-[2-(4-
Nitro-phenyl)-vinyl]-phenylamine (4). To a solution of compound
2 (0.10 g, 0.4mmol) in THF (20 mL) was added SnCl2 (1.50 g)
dissolved in 1N HCl (10 mL). The reaction mixture was stirred
overnight at room temperature. The acidic solution was then
neutralized using 1N NaOH and extracted with ethyl acetate (3 ×
20 mL). The combined organic phases were washed with water
and brine, dried over Na2SO4, and concentrated. Purification with
flash column (HE:EA ) 2:1 to 1:1) yielded 4,4′-diamino-trans-
stilbene (3, 0.03 g, 40%) and 4-[2-(4-nitro-phenyl)-vinyl]-pheny-
lamine (4). 1H NMR of 3 (300 MHz, CDCl3): 7.31 (d, J ) 8.4 Hz,
4 H), 6.86 (s, 2 H), 6.68 (d, J ) 8.4 Hz, 4 H). HR-ESIMS of 3:
m/z calcd for C14H14N2 (M + H+): 211.1230, found 211.1225.
1
Melting point of 3: 206.1-207.3 °C. H NMR of 4 (300 MHz,
CDCl3): 8.22 (d, J ) 8.0 Hz, 2 H), 7.59 (d, J ) 6.86 Hz, 2 H),
7.41 (d, J ) 8.57 Hz, 4 H), 7.22 (d, J ) 17.14 Hz, 1 H), 6.97 (d,
J ) 12.57 Hz, 1 H), 6.72 (d, J ) 10 Hz, 2 H).
1.8. Synthesis of 4-[2-(4-Tributylstannanyl-phenyl)-vinyl]-phe-
nylamine (10). Under Ar, the substrate 9 (0.05 g, 0.15 mmol) was
mixed with (Bu3Sn)2 (1 mL), Pd(PPh3)4 (0.02 g), and Et3N (5 mL).
The mixture was sealed in a vial and heated to 80 °C for 1 day.
The solvent was evaporated in vacuum, and the residue was purified
1.3. Synthesis of Dimethyl-{4-[2-(4-nitro-phenyl)-vinyl]-phenyl}-
amine (5). To a solution of 4-dimethylamino-benzaldehyde (2.24
g, 15 mmol) and (4-nitro-benzyl)-phosphoric acid diethyl ester (1,
4.10 g, 15 mmol) in DMF (20 mL) and EtOH (20 mL) was added
to NaOCH3 (1.62 g, 30 mmol). The suspension was stirred and
refluxed for 3 h. After cooled to room temperature, the precipitate
was filtered and washed thoroughly with ethanol to give dimethyl-
{4-[2-(4-nitro-phenyl)-vinyl]-phenyl}-amine (5, 2.55 g, 65%) as red
solid. 5 was used without further purification.
1.4. Synthesis of Dimethyl-{4-[2-(4-amino-phenyl)-vinyl]-phen-
yl}-amine (6). To a solution of 5 (2.55 g, 9.5mmol) in EtOH (100
mL) was added to SnCl2 (8.58 g, 38 mmol). The resulting mixture
was refluxed for 4 h. The solvent was then removed under vacuum
and NaOH (2 mol/L, 40 mL) was added to the residue. The crude
solid was filtered and suspended in ethyl acetate (200 mL). The
precipitates were then filtered to give dimethyl-{4-[2-(4-amino-
1
by column to give 10 (44 mg, 0.09 mmol, yield: 60%). H NMR
(400 MHz, CDCl3): 7.42 (s, 4 H), 7.33 (d, J ) 7.87 Hz, 2 H), 7.03
(d, J ) 16.18 Hz, 1 H), 6.89 (d, J ) 16.24 Hz, 1 H), 6.66 (d, J )
8.08 Hz, 2 H), 1.55 (m, 6 H), 1.34 (m, 6 H), 1.05 (t, 6 H), 0.89 (t,
9 H).
1.9. Radiosynthesis of 4-[2-(4-[125I]Iodo-phenyl)-vinyl]-pheny-
lamine ([125I]9). To a sealed vial were added 10 (50 µL, 50 µg in
50 µL of ethanol), [125I] sodium iodide, and 1N HCl (100 µL).
Subsequently, 100 µL of H2O2 (3%, in water) was added via a
syringe at room temperature. After 10 min, the iodination reaction
was terminated by an addition of saturated NaHSO3, and the
resulting solution was neutralized to pH 7-8 by adding a saturated
NaHCO3 solution. The mixture was extracted with ethyl acetate (3
× 1 mL). The combined organic layers were dried over Na2SO4,
and the solvent was removed by a stream of dry nitrogen gas. The
residue was purified by high performance liquid chromatography
(HPLC; C-18 column; acetonitrile; DMGA (5 mM, pH 7.4): 60/
40; flow rate: 1 mL/min; retention time: 21 min) to get 18.5 MBq
of final pure product with radiochemical purity over 98% and a
1
phenyl)-vinyl]-phenyl}-amine (6, 1.45 g, 64%) as gray solid. H
NMR (300 MHz, CDCl3): 7.40 (d, J ) 8.62 Hz, 2 H), 7.33 (d, J )
8.40 Hz,2 H), 6.86 (d, J ) 5.92 Hz, 2 H), 6.76 (d, J ) 8.34 Hz, 2
H), 6.69 (d, J ) 8.22 Hz, 2 H), 2.99 (s, 6 H). HR-ESIMS: m/z
calcd for C16H18N2 (M + H+), 239.1543; found, 239.1542. Melting
point: 167.7-168.5 °C.