H. Li, X. Zhou, H. Ye, X. Sun, and P. Zhang
Vol 000
inflammatory infiltration, essentially in line with our
observations regarding the clinical symptoms of EAE.
To our knowledge, the expression of ROCK kinase
inhibitor is significantly upregulated in EAE mice, with
symptoms of EAE improving with administration of
fasudil. This may relate to the inhibition of ROCK-II
and the decrease of inflammatory infiltration [20,21];
Zhang et al. reported that fasudil oral delays the onset
of EAE, slows down EAE symptoms, reduces weight
loss, and reduces EAE myelin loss and CNS
The synthesis of compounds 1 and 2 has been shown in
our previous experiment [13], not repeated here.
6H-1-(5-Isoquinolinesulfonyl)-4-(β-hydroxy
ethyl)-1,4-
diazepine (compound 3). Fasudil hydrochloride (1.00 g,
2.74 mmol) was slowly added to a saturated sodium
bicarbonate solution (0.52 g, 4.93 mmol). The mixture
kept its pH at a constant value (pH 5–6). This solution
was stirred for 30 min and extracted with 20-mL CH2Cl2.
Then, it was dried over anhydrous MgSO4, followed by
filtration and concentration. The residue dissolved in 20-
mL CH2Cl2 was dropwise added to triethylamine (0.33 g,
3.30 mmol). Then, 2-bromoethanol (0.90 g, 8.23 mmol)
was added slowly to the mixture. The reaction was
allowed to develop for 30 h at room temperature. And
then, the organic layer was concentrated. The residue was
purified by flash chromatography (ethyl acetate) to afford
compound 3 (0.57 g, 65%) as a yellow oil. 1H NMR
(500 MHz, CDCl3) δ: 1.81–1.86 (m, 2H), 2.61 (t,
J1 = 6.0 Hz, J2 = 6.0 Hz, 2H), 2.65–2.68 (m, 4H), 3.47–
3.51 (m, 4H), 3.59 (m, 2H), 7.65–7.68 (m, 1H), 8.17 (d,
J = 9.0 Hz, 1H), 8.34 (d, J = 9.0 Hz, 1H), 8.45 (d,
J = 6.0 Hz, 1H), 8.66 (d, J = 6.0 Hz, 1H), 9.33 (s, 1H).
13C NMR (125 MHz, CDCl3) δ: 8.50, 18.50, 45.67,
45.76, 55.19, 55.97, 57.69, 116.88, 126.62, 128.84,
130.60, 131.69, 132.79, 133.01, 134.02, 145.02, 153.55.
HRMS (ESI), Calcd C16H21N3O3S: [M + H] + m/z:
336.1374. Found: 336.1376. IR (KBr cmÀ1): 3489, 3419,
2974, 2737, 1366, 1147, 1074, 678, 570.
inflammation infiltration. Fasudil oral has
a good
therapeutic effect and can significantly reduce the core
of the CNS, which can significantly alleviate the loss of
myelin and inflammatory lesions in the CNS [22].
Compounds 1, 2, and 6, as the novel ROCK inhibitors,
might provide a therapeutic potential against injuries to
the human CNS.
CONCLUSION
Compounds 1 and 2 displayed excellent Rho kinase
inhibitory activity; we designed and synthesized their
enantiomers to reveal the inhibitory effects of chirality on
Rho kinase. It is found that their enantiomers also
exhibited much better Rho kinase inhibitory activity and
strongly promoted synapse formation than fasudil.
Establishing the EAE model, after added antigens, the
changes of limbs and tail were observed and scored by
clinical evaluation. The synthetic compounds may
simultaneously reduce symptoms of EAE and inhibit
inflammatory infiltration of the CNS. Thus, compounds
1, 2, 5, and 6 may be potential candidates for inhibition
of Rho kinase and should be considered for further
experimental study in relation to MS. Further studies on
combination mode between inhibitors and Rho kinase are
currently under way.
4-(Isoquine-5-sulfonyl)-[1,4]diazepane-1-carboxylic
tert-butyl ester) (compound 4).
acid
Fasudil hydrochloride
(5.00 g, 14 mmol) was slowly added to a saturated
sodium bicarbonate solution (2.62 g, 24 mmol). The
mixture kept its pH at a constant value (pH 5–6). The
mixture was stirred 0.5 h and extracted with 50-mL
CH2Cl2 for three times. The organic layers were dried
and concentrated. The residue dissolved in 50-mL
CH2Cl2 was added dropwise to the flask containing Di-
tert-butylpyrocarbonate (4.49 g, 20.6 mmol) and
triethylamine (1.39 g, 13.7 mmol). The mixture was
stirred at 0–5°C for 2 h. After dried and concentrated,
the residue was purified to afford compound 4 (5.25 g,
EXPERIMENTAL
1
97%) as a yellow oil. H NMR (400 MHz, CDCl3) δ:
General.
All the chemicals were purchased from
8.84 (s, 1H), 8.53 (d, J = 10 Hz, 1H), 8.25 (d,
J = 5 Hz, 1H), 8.05–8.10 (m, 1H), 7.91 (d, J = 10 Hz,
1H), 7.67 (d, J = 15 Hz, 1H), 3.49–3.58 (m, 4H), 3.35–
3.43 (m, 4H), 1.43(s, 9H). 13C NMR (100 MHz,
CD3OD) δ: 27.12, 28.76, 40.10, 42.25, 43.92, 47.07,
70.97, 120.27, 127.35, 127.10, 130.57, 132.54, 137.17,
.51, 136.13, 142.71, 152.20, 155.79. HRMS (ESI),
Calcd C19H25N3O4S: [M + H] + m/z: 392.4814. Found:
392.4815. IR (KBr cmÀ1): 3019, 2479, 1647, 1480,
Guangfu Technology Development Co., Ltd., Tianjin
(China) and Chase Sun Pharmaceutical Co., Ltd., Tianjin
(China) unless otherwise stated. All reagents were used
as purchased from commercial suppliers without further
purification. Solvents were dried and purified according
to standard procedures before use. The course of reactions
was monitored by thin-layer chromatography (silica gel
GF254s); flash chromatography was performed using 200–
300 mesh silica gel. 1H and 13C NMR spectra were recorded
on INOVA400/600 Hz spectrometer with tetramethylsilane
as an internal standard. HRMS was recorded on
MicroOTOF-Q II (CycLex Co., Ltd, Nagano, Japan).
1330, 1160, 724, 582.
(S)-6H-1-(5-Isoquinolinesulfonyl)-2-hydroxymethyl-1-
pyrrolidine (compound 5). Isoquinoline sulfonyl chloride
hydrochloride (5 g, 18.9 mmol) was slowly added to a
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet