Synthesis and biological evaluation of lipid-soluble prodrugs of anethole dithiolthione

Article abstract of DOI:10.1016/j.cclet.2010.07.014

16 ADT carboxylate esters were prepared by means of esterification and these compounds were expected to increase the bioavailability of 4-hydroxyanehole trithione. In vivo studies showed that ADT concentration of 3a in plasma was much higher than that of ATT during 120 min. Compound 3a could reach blood peak values of ADT at 660.6 ng/mL which was about 14 times of that by ATT. Additionally, the acute toxicity assay indicated high safety of compound 3a that the maximum tolerated dose was no less than 3.25 g/kg.

Full text of DOI:10.1016/j.cclet.2010.07.014

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Chinese Chemical Letters 21 (2010) 1427–1429
www.elsevier.com/locate/cclet
Synthesis and biological evaluation of lipid-soluble prodrugs
of anethole dithiolthione
Mei Guan a, Wei Fan b, Shan Qian b, Rui Qi Xiao b, Yong Wu b,
*
a West China Hospital, Sichuan University, Chengdu 610041, China
b Key Laboratory of Drug Targeting of China Education Ministry, West China School of Pharmacy, Sichuan University,
Chengdu 610041, China
Received 23 March 2010
Abstract
16 ADT carboxylate esters were prepared by means of esterification and these compounds were expected to increase the
bioavailability of 4-hydroxyanehole trithione. In vivo studies showed that ADT concentration of 3a in plasma was much higher than
that of ATT during 120 min. Compound 3a could reach blood peak values of ADTat 660.6 ng/mL which was about 14 times of that
by ATT. Additionally, the acute toxicity assay indicated high safety of compound 3a that the maximum tolerated dose was no less
than 3.25 g/kg.
# 2010 Yong Wu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: Synthesis; Anethole dithiolthione; Liposoluble; Bioavailability; Prodrug
Anethol trithione (ATT), carrying trithione structure, is a relatively new chologogue. ATT and its analogues have
been studied as sialagogue, antischistosomial, choleretic and hepatic protectant [1,2], which are widely used in clinical
for treating symptoms associated with hepatobiliary dysfunctions without any major adverse reactions being noted [3].
ATT can increase human cholinergic, adrenergic responsiveness and prevent the up-regulation in muscarinic
acethylcholine receptor density. It is also a potent inducer of phase II enzymatic detoxification systems and can
increase the level of tissue’s glutathione (GSH) which can deactivate many chemicals by conjugating them [4,5].
Besides the above effects, it was found that ATT analogues could prevent cellular superoxidant damages and
carcinogenesis produced by certain compounds [6].
The previous research [7] described that ATT was firstly metabolized into 4-hydroxyanehole trithione (ADT) in
plasma and then ADTwas conjugated with sulfuric acid or glycuronic acid to excrete. The primary pharmacodynamic
active constituent was not ATT itself but its demethyl metabolite ADT, since ATT lose methyl group in liver after
administration. However, it must be noted that the metabolizing rate of ATTwas comparatively low which limited the
pharmacodynamic effect of ADT. Inspired by those previous research results and considering the active constituent
ADT structural characteristics, 16 ADT carboxylate esters (Fig. 1) were prepared by means of esterification as
prodrugs of ADT, which was capable of improving the biological availability of ADT.
* Corresponding author.
E-mail address: wyong@scu.edu.cn (Y. Wu).
1001-8417/$ – see front matter # 2010 Yong Wu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
doi:10.1016/j.cclet.2010.07.014

1[(ig._1)TD$FIG] 428
M. Guan et al. / Chinese Chemical Letters 21 (2010) 1427–1429
Fig. 1. ATT, ADT and ADT carboxylate esters 3a–3p.
[(Schme_1)TD$FIG]
Scheme 1. The synthetic route of 3a–3p. Condition and regent: (a) pyridine hydrochloride, 215 8C, 3 h; (b) DCC, DMAP, CH2Cl2, r.t.
In order to improve the fat-solubility of ADT, the methyl group was substituted by various lipid groups via ester
linkage with hydroxyl group of ADT. These compounds were prepared by the procedures illustrated in Scheme 1. We
managed to afford ADT (2) with satisfied yield (84.5%) by treating ATT with pyridine hydrochloride under N2
atmosphere at 215 8C. To a solution of 2 (1.00 mmol) in CH2Cl2, DCC (1.07 mmol), DMAP (0.15 mmol) and R-
carboxylic acid (0.97 mmol) were added. The mixture was stirred at room temperature until the 4-hydroxyanehole
trithione (2) was consumed nearly (monitored by TLC). The filtrate was concentrated and purified by column
chromatography. In total, 16 new compounds 3a–3p were prepared, their structures were confirmed by IR, 1H NMR,
ESI-MS [8].
In the present works, we chose compound 3a for bioavailability text. Freshly prepared solutions of the compound in
peanut oil were given by intra-gastric injection in a dose of 250 mg/kg body weight of female mice. At specific time
intervals the animals were killed, and blood samples were collected for pretreatment and analyzed by a HPLC
determination method which was established and applied to investigate the major metabolite (ADT) of this compound.
The result was shown the compound could not be detected at the first time interval (5 min), which may be caused by the
high metabolic rate of them in plasma. It was obvious that the test compound could reach the peak values of blood drug
concentration (660.6 ng/mL) of ADT at 60 min which was much higher and more swiftly than the concentration
(48.4 ng/mL) of ATTat 120 min. This can be explained by the better lipid-solubility of compound 3a than ATT, which
can enhance the uptake rate of the compound in mice intestinal tract. Moreover, the ADT concentrations of compound
3a in plasma were remarkably higher than that of ATT during 180 min as expect. The investigation results are listed in
Table 1.
Since the good bioavailability of compound 3a, the safety of this compound is a non-negligible factor. We
conducted the acute toxicity assay of this compound with mice. The maximum tolerated dose (MTD) of compound 3a
Table 1
The comparison of plasma concentrations of ADT following i.g. administration of ATT and compound 3a in mice (ng/mL).
Compounds
Time (min)
5
10
15
30
45
60
120
180
3a
ATT
65.3
16.0
101.2
21.4
178.3
29.1
145.8
33.4
243.2
37.2
660.6
46.7
49.8
48.4
nd
45.0
nd: not detected.

M. Guan et al. / Chinese Chemical Letters 21 (2010) 1427–1429
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was no less than 3.25 g/kg while the value of positive control ATT was 0.85 g/kg which indicated that the test
compound was more safe than ATT.
As all above shown, compound 3a is a potential hepatic protectant which exhibit better bioavailability and potency
than ATT. The bioavailability and potency evaluation of other synthesized compounds is underway.
References
[1] C.K. Sen, Biochem. Pharmacol. 55 (1998) 1747.
[2] S.S. Ansher, P. Dolan, E. Bueding, Food Chem. Toxicol. 24 (1986) 405.
[3] P.A. Egner, T.W. Kensler, P.M. Dolan, et al. Carcinogenesis 15 (1994) 177.
[4] C.M. Odile, Methods Enzymol. 252 (1995) 316.
[5] T. Nagano, M. Takeyama, J. Pharm. Pharmacol. 53 (2001) 1697.
[6] Z. Yusheng, B.G. Gary, Mol. Cancer Ther. 3 (2004) 885.
[7] R. Gmelin, F. Lagler, Azneim. Forsch. 13 (1963) 130.
[8] Select data of compound 3. 3a: 4-(3-Thioxo-3H-1,2-dithiol-5-yl)phenyl acetate: yield 82.7%; Mp 140–142 8C; IR (film, cmÀ1): n 3021, 2922,
1763, 1363, 1185, 1023; 1H NMR (400 MHz, CDCl3): d 7.69 (dt, 2H, J1 = 8.8 Hz, J2 = 2.0 Hz), 7.40 (s, 1H), 7.24 (dt, 2H, J1 = 8.8 Hz,
J2 = 2.0 Hz), 2.34 (s, 3H); ESI-MS (m/z): 269.0 ([M+H]+). 3d: 4-(3-Thioxo-3H-1,2-dithiol-5-yl)phenyl isobutyrate: yield 97.5%; Mp 99–
100 8C; IR(film, cmÀ1): n 3046, 2977, 2932, 1748, 1386, 1164, 1027; 1H NMR (400 MHz, CDCl3): d 7.67 (dt, 2H, J1 = 8.8 Hz, J2 = 2.0 Hz), 7.41
(s, 1H), 7.22 (dt, 2H, J1 = 8.8 Hz, J2 = 2.0 Hz), 2.84 (m, 1H), 1.33 (d, 6H, J = 7.2 Hz); ESI-MS (m/z): 297.1 ([M+H]+). 3f: 4-(3-Thioxo-3H-1,2-
dithiol-5-yl)phenyl 2-phenylacetate: yield 85.9%; Mp 134–136 8C; IR (film, cmÀ1): n 3028, 2914, 1353, 1231, 1169, 1026; 1H NMR (400 MHz,
CDCl3): d 7.65 (d, 2H, J = 8.4 Hz), 7.38 (d, 5H, J = 4 Hz), 7.25 (s, 1H), 7.20 (d, dH, J = 8.4 Hz), 3.88 (s, 2H); ESI-MS (m/z): 345.0 ([M+H]+). 3i:
4-(3-Thioxo-3H-1,2-dithiol-5-yl)phenyl cinnamate: yield 91.2%; Mp 161–162 8C; IR (film, cmÀ1): n 3059, 1729, 1630, 1309, 1189, 1025; 1H
NMR (400 MHz, CDCl3): d 7.90 (d, 1H, J = 16 Hz), 7.71 (dt, 2H, J1 = 8.8 Hz, J2 = 2.0 Hz), 7.60 (m, 2H), 7.44 (m, 3H), 7.42 (s, 1H), 7.33 (dt,
2H, J1 = 8.8 Hz, J2 = 2.0 Hz), 6.62 (d, 1H, J = 16 Hz); ESI-MS (m/z): 357.3 ([M + H]+). 3p 4-(3-Thioxo-3H-1,2-dithiol-5-yl)phenyl-5,6,7,8-
tetrahydronaphthalene-1-carboxylate: yield 97.8%; Mp 114–115 8C; IR (film, cmÀ1): n 3026, 2930, 1757, 1598,1220, 1028; 1H NMR
(400 MHz, CDCl3): d 7.96 (m, 1H), 7.74 (dt, 2H, J1 = 8.8 Hz, J2 = 2.0 Hz), 7.44 (s, 1H), 7.36 (dt, 2H, J1 = 8.8 Hz, J2 = 2.0 Hz), 7.34 (b, 1H),
7.23 (d, 2H, J = 7.6 Hz), 3.15 (m, 2H), 2.87 (m, 2H), 1.85 (m, 4H); ESI-MS (m/z): 385.4 ([M+H]+).