G Model
JPC 10105 No. of Pages 9
S. Hikage et al. / Journal of Photochemistry and Photobiology A: Chemistry xxx (2015) xxx–xxx
7
Therefore, we concluded that not only the function of the caged
ASO containing thiochromone-type PLPG was recovered by
photoirradiation, but also the photodeprotection process could
be monitored by fluorescence measurements, which cannot be
accomplished by typical PLPGs, in vitro experiment.
light (Blak-ray B-100SP (365 nm: 111
m
W/cmꢁ2); UVP LCC), LED
lamp (SDL-445-LM-030T (445 nm: 2.39
Dream Laser Technology Co., Ltd.), and xenon lamp (MAX-303
(330 nm: 1.02
W/cmꢁ2); ASAHI SPECTRA).
m
W/cmꢁ2); Shanghai
m
4.2. Compound 3
3. Conclusion
Under N2 atmosphere, 4-dimethylaminopyridine (DMAP)
(8 mg, 0.064 mmol), 2,4,6-triisopropyl benzenesulfonyl chloride
(TPBS) (117 mg, 0.385 mmol) were added to a solution of acetyl
thymidine (1) (100 mg, 0.321 mmol) in dry acetonitrile, then the
resulted solution was stirred at 45 ꢀC for 4 h. To this solution was
added 2-hydroxymethyl-3-phenyl thiochromone S,S-dioxide (2)
(96 mg, 0.321 mmol), and then the resulted solution was stirred at
the same temperature for 12 h. The reaction mixture was extracted
with chloroform, and the organic layer was washed with 1 N aq.
HCl and saturated aq. NaHCO3. Then, the collected organic layer
was dried over sodium sulfate followed by concentration in vacuo
and silica gel column chromatography (ethyl acetate/hexane = 2/1)
to afford target compound 3 (27% yield) as a yellow solid: Rf value
0.24 (ethyl acetate/hexane = 1/1); IR (neat) nmax 3648, 3086, 3063,
3016, 2950, 2359, 2339, 1743, 1713, 1661, 1464, 1373, 1307, 1231,
To monitor the photodeprotection mechanism of caged
compounds by fluorescence measurements, a novel caged nucleic
acid with thiochromone-type PLPG was synthesized. This product
showed a unique property: Highly fluorescent byproducts were
obtained during photodeprotection. As expected, when the caged
nucleic acid was photoirradiated, fluorescent byproducts were
formed. However, they were different from the expected
fluorophore, and the original nucleic acid was not recovered
during a short photoirradiation. The original nucleic acid was
successfully recovered after a longer photoirradiation. The photo-
deprotection process was monitored by the fluorescence measure-
ments of a sample solution during a prolonged irradiation. A novel
caged ASO containing thiochromone-type PLPGs was synthesized
to control the bioactivity by photoirradiation. The fluorescence
peak pattern of the caged ASO solution was similar to that of the
caged nucleic acid. The original ASO was successfully recovered by
appropriate photoirradiation based on the fluorescence measure-
ments as confirmed by a bioactivity assay, although the photo-
deprotection efficiency of the novel caged nucleic acid is low
compared to typical PLPGs. Studies on more efficient photo-
deprotection of caged compounds containing thiochromone-type
PLPGs are underway.
1059, 757 cmꢁ1; 1H NMR (CDCl3)
d: 8.16 (dd, J = 1.0, 8.0 Hz,1H), 8.01
(d, J = 8 Hz, 1H), 7.84 (dt, J = 1.0, 7.5 Hz, 1H), 7.74 (dt, J = 1.5, 8.0 Hz,
1H), 7.42 (m, 3H), 7.30 (d, J = 7.0 Hz, 2H), 7.18 (d, J = 1.0 Hz, 1H), 6.28
(dd, J = 5.0, 8.5 Hz, 1H), 5.21 (dd, J = 17.0, 60.0 Hz, 2H), 5.19 (m, 1H),
4.34 (m, 2H), 4.24 (dd, J = 3.5, 5.5 Hz, 1H), 2.50 (ddd, J = 2.0, 5.8,
14.0 Hz, 1H), 2.13 (m, 1H), 2.11 (s, 3H), 2.09 (s, 3H), 1.89 (d, J = 1.0 Hz,
3H); 13C NMR (CDCl3)
d: 178.1,170.5,170.3,162.6,150.3,146.2,141.7,
141.1, 134.6, 133.3, 133.0, 131.7, 129.3, 129.0, 128.9, 128.8, 128.5,
123.0, 110.3, 85.9, 82.4, 74.4, 64.0, 38.7, 37.9, 21.0, 21.0, 13.5; HRMS
(ESI) Calcd for C30H28N2NaO10
631.1362.
4. Experimental
S
[M + Na]+: 631.1362; Found
4.1. General
Most commercially available reagents were used without
further purification. Spectrograde solvents purchased from Wako
pure chemical industries were used for the photoreaction.
Dehydrate solvents purchased by Wako pure chemical industries
and Kanto Chemical Co. Inc., were used for the organic synthesis.
1H NMR and 13C NMR spectra were recorded on a JEOL JNM-
ECP500 (1H at 500 MHz, 13C at 126 MHz). 1H NMR spectra are
reported as a chemical shift in ppm based on the peak of CDCl3
4.3. Fluorescent byproducts (5 and 6; absolute configuration is
unknown)
5 (white solid); IR (neat) nmax 3525, 2925, 2360, 1744, 1704,
1648, 1465, 1452, 1313, 1230, 1192, 1107, 1044, 754 cmꢁ1 1H NMR
;
(CDCl3) d: 8.08 (dd, J = 1.0, 8.0 Hz, 1H), 7.97 (d, J = 7.5 Hz, 1H), 7.75
(dt, J = 1.0, 8.0 Hz, 1H), 7.60 (dt, J = 1.0, 8.0 Hz, 1H), 7.54 (d, J = 8.0 Hz,
1H), 7.38 (d, J = 8.0 Hz, 1H), 7.33 (dt, J = 7.5, 1.5 Hz, 1H), 7.20 (t,
J = 7.0 Hz, 1H), 7.11 (d, J = 1.5 Hz, 1H), 6.06 (t, J = 6.5 Hz, 1H), 5.27 (dd,
J = 7.5, 8.5 Hz, 1H), 5.08 (m, 1H), 4.55 (dd, J = 7.5, 13.5 Hz, 1H), 4.36
(dd, J = 9.0, 13.5 Hz, 1H), 4.29 (s, 1H), 4.28 (s, 1H), 4.19 (dd, J = 4.0,
6.0 Hz, 1H), 2.13 (m, 1H), 2.11 (s, 6H), 1.87 (s, 3H); 13C NMR (CDCl3)
(d= 7.26 ppm), integration, multiplicity (s = singlet, d = doublet,
t = triplet, q = quartet, dd = doublet of doublet, ddd = doublet of
doublet of doublets, dt = doublet of triplet, m = multiplet), and the
coupling constant (Hz). 13C NMR spectra are reported as chemical
shifts in ppm based on the middle peak of CDCl3 (d= 77.0 ppm). UV/
d
: 170.5, 170.3, 163.1, 154.7, 150.5, 147.1, 134.1, 133.1, 129.8, 127.6,
vis spectra and Fluorescence spectra were measured in a quartz
cell (10 mm thickness) on a JASCO V-630 spectrometer and FP-
6500 spectrometer respectively. ESI-MS spectra were performed
with a JEOL JMS-T100LC. Infrared spectra were measured on a
JASCO FT/IR-4200 spectrometer. Luminescence measurement was
performed on a Molecular Devices SpectraMax L microplate reader.
Analytical high-performance liquid chromatography (HPLC) was
carried out using Hitachi L-2300 (Column Oven), L-2130 (Pump),
and L-2400 (UV–vis Detector) instruments with a Tokyo Chemical
Industry Co., Ltd. (TCI) LC-ODS 2000 column. Preparative HPLC was
carried out using Hitachi L-7150 (Pump), and L-7420 (UV–vis
Detector) instruments with a TCI LC-ODS 2000 column. The HPLC
analyses were run at 25 ꢀC and eluted with a 1:1 water/acetonitrile
mixture at a flow rate of 0.5 mL/min under an isocratic condition
(UV detected at 333 nm). The isolation of new fluorophore by HPLC
were run at 25 ꢀC and eluted with a 61:39 water/acetonitrile
mixture at a flow rate of 10 mL/min under an isocratic condition
(UV detected at 333 nm). The light sources were follows: the black
126.9, 126.2, 125.6, 123.9, 123.7, 119.1, 112.0, 111.3, 110.4, 85.9, 82.4,
74.1, 63.9, 54.7, 39.9, 37.4, 21.1, 21.0, 13.4; HRMS (ESI) Calcd for
C
30H28N2O10S [M + Na]: 631.1362; Found 631.1360.
6 (white solid); IR (neat) nmax 3628, 3019, 2930, 2360, 1743,
1706, 1649, 1465, 1452, 1313, 1231, 1140, 1107, 753 cmꢁ1 1H NMR
;
(CDCl3) d: 8.05 (d, J = 8.0 Hz, 1H), 7.96 (d, J = 8.5 Hz, 1H), 7.76 (dt,
J = 1.0, 7.5 Hz, 1H), 7.59 (dt, J = 1.5, 11.0 Hz, 1H), 7.55 (dd, J = 2.5,
6.0 Hz, 2H), 7.35 (dt, J = 1.5, 8.5 Hz,1H), 7,24 (t, J = 7.5 Hz,1H), 7.17 (d,
J = 1.5 Hz, 1H), 6.14 (dd, J = 5.5, 8.0 Hz, 1H), 5.28 (dd, J = 7.8, 8.8 Hz,
1H), 5.20 (m, 1H), 4.54 (dd, J = 8.5, 14.0 Hz, 1H), 4.34 (d, J = 4.0 Hz,
2H), 4.32 (dd, J = 8.5, 14.0 Hz, 1H), 4.24 (q, J = 3.0 Hz, 1H), 2.52 (ddd,
J = 1.5, 5.8,14.3 Hz,1H), 2.14 (m,1H), 2.13 (s, 3H), 2.10 (s, 3H),1.84 (d,
J = 1.0 Hz, 3H); 13C NMR (CDCl3)
d: 170.6, 170.4, 163.1, 154.8, 150.7,
147.0, 134.0, 133.2, 133.0, 129.8, 127.4, 126.9, 126.3, 125.6, 124.0,
123.7, 119.3, 112.1, 111.4, 110.3, 86.3, 82.6, 74.3, 63.9, 54.8, 39.8, 37.9,
21.1, 21.0, 13.4; HRMS (ESI) Calcd for C30H28N2O10S [M + Na]:
631.1362; Found 631.1367.
Please cite this article in press as: S. Hikage, et al., Synthesis of novel caged antisense oligonucleotides with fluorescence property, J.