Therapeutic effects of isothiocyanate prodrugs on rheumatoid arthritis

Article abstract of DOI:10.1016/j.bmcl.2018.01.009

Isothiocyanates 7a and 7b have poor stability and aqueous solubility. To address these problems, prodrugs 8a and 8b were synthesized. Prodrugs 8a and 8b were stable in HEPES buffer at pH 4.4, but released the active compounds 7a and 7b in HEPES buffer at

Full text of DOI:10.1016/j.bmcl.2018.01.009

Bioorganic & Medicinal Chemistry Letters xxx (2018) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Therapeutic effects of isothiocyanate prodrugs on rheumatoid arthritis
a,
Yin Jiang a,b,c, Hui-Ying Li b,c, Xiao-Hui Li a,b, Jun Lu a,b, Quan Zhang a, Cui-Gai Bai b, , Yue Chen
⇑
⇑
a The State Key Laboratory of Medicinal Chemical Biology, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), College of Pharmacy,
and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300353, PR China
b High-throughput Molecular Drug Discovery Center, Tianjin International Joint Academy of BioMedicine, Tianjin 300457, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Isothiocyanates 7a and 7b have poor stability and aqueous solubility. To address these problems, pro-
drugs 8a and 8b were synthesized. Prodrugs 8a and 8b were stable in HEPES buffer at pH 4.4, but released
the active compounds 7a and 7b in HEPES buffer at pH 7.4 and in mouse plasma, respectively. Compound
8a and especially compound 8b showed anti-inflammatory effects. Compound 8b demonstrated signifi-
cant efficacy in animal models of traumatic inflammation, acute inflammation and rheumatoid arthritis.
Compound 8b also did not cause appreciable toxicity in mice after 5 weeks at a daily dose of 200 mg/kg.
Ó 2018 Published by Elsevier Ltd.
Received 22 November 2017
Revised 5 January 2018
Accepted 7 January 2018
Available online xxxx
Keywords:
Isothiocyanate prodrug
Traumatic inflammation
Acute inflammation
Rheumatoid arthritis
Active drug
Rheumatoid arthritis (RA) is
a
heterogeneity, chronic,
dimethylamine. Treatment of compound 3 with SOCl2 then gave
compound 4, which was treated with potassium thioacetate to
provide compound 5. Hydrolysis of compound 5 in methanol con-
taining hydrazine hydrate then afforded compound 6. Treatment of
isothiocyanates 7a and 7b with compound 6 gave prodrugs 8a and
8b, respectively (Scheme 1).
Transformation from prodrugs 8a and 8b to active drug was
determined in HEPES buffer and plasma to simulate stomach,
intestines and blood environment in vivo. In HEPES buffer at
pH 4.4, compounds 8a and 8b were stable, and no significant
amount of 7a or 7b was released within 24 h (Fig. 1). In
HEPES buffer at pH 7.4, compounds 8a and 8b were slowly
converted to the active drugs, 7a and 7b. Compound 8a
achieved balance at 1 h and then continued to transform to
7a. Compound 8b achieved balance at 2 h and then remained
unchanged (Fig. 2). In plasma, compounds 8a and 8b were
transformed to 7a and 7b, and had almost completely decom-
posed after 24 h.
systemic autoimmune diseases, characterized by symmetrical
polyarthritis. Women are approximately three times more likely
to develop RA than men.1 Although the exact etiology of RA is
unclear, it is generally believed that this is a genetic disease dri-
ven by multiple factors. Individuals with a family history of RA
have a 3–9-fold higher risk of developing the disease than the
general population.2 The collagen-induced arthritis model3,4
and the adjuvant arthritis model5,6 are the most widely used
animal models of RA.
Isothiocyanate compounds, such as sulforaphane (SFN), erucin,
phenethyl isothiocyanate and benzyl isothiocyanate, are abound
in plants, especially cruciferous plants. Previous studies have
shown that isothiocyanate compounds have anti-inflammatory
effect, represented by SFN.7–15 But isothiocyanates have poor sta-
bility and solubility, for example, the half-life of SFN is 2.2 h in
rats16 and 1.77 0.13 h in one human study.17
Herein, 8a and 8b, prodrugs of isothiocyanates 7a and 7b, were
synthesized and evaluated for stability and efficacy. Preliminary
toxicity data was also presented.
Triethylene glycol was treated with 4-dimethylaminopyridine
(DMAP) and p-toluenesulfonyl chloride (PTSC) to obtain compound
2, which was further converted to compound 3 in aqueous
In mouse plasma, compounds 8a and 8b were also slowly con-
verted to the active drugs, 7a and 7b. Tmax were 2 h. Compounds
8a and 8b almost completely decomposed in 24 h and 10 h, respec-
tively. The half-lives of compounds 8a and 8b in mouse plasma
were about 6 h in vitro, so compounds 8a and 8b were thus likely
to be stable in vivo (Fig. 3).
Pharmacodynamics of the isothiocyanate prodrugs were evalu-
ated by the neutrophilic inflammation model in EGFP transgenic
zebrafishes, the carrageenan-induced paw edema model in rats
⇑
Corresponding authors.
E-mail addresses: baicuigai@tjab.org (C.-G. Bai), yuechen@nankai.edu.cn
(Y. Chen).
c
These authors contributed equally to this work.
https://doi.org/10.1016/j.bmcl.2018.01.009
0960-894X/Ó 2018 Published by Elsevier Ltd.
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Y. Jiang et al. / Bioorganic & Medicinal Chemistry Letters xxx (2018) xxx–xxx
Scheme 1. Synthesis of isothiocyanate prodrugs 8a–b. Reagents and conditions: (i) DMAP, PTSC, Et3N, THF, 0 °C to r.t, 44.4%; (ii) dimethylamine aqueous, r.t, 62.9%; (iii) SOCl2,
CHCl3, 0 °C to 70 °C, 95%; (iv) Potassium thioacetate, DMF, 75 °C, 62.6%; (v) Hydrazine hydrate, MeOH, r.t, Ar, 51.1%; (vi) DCM, Et3N, r.t, 64.5% for 8a and 70.3% for 8b.
Fig. 1. Concentration–time curve of 8a (d), 7a (j), 8b (▲), and 7b (.) in HEPES buffer of pH 4.4.
and the Freund ’s complete adjuvant model in rats. These models
were traumatic inflammation, acute inflammation and RA,
respectively.
Dexamethasone (DEX) and methotrexate (MTX) were used as
positive controls. (Figs. 4–6). The animal experiments have been
approved by ethics committee of Tianjin International Joint Acad-
emy of Biomedicine.
In the zebrafish neutrophilic inflammation model, the num-
ber of migrating neutrophils is an important evaluation index,
with fewer migrating cells indicating better efficacy (Fig. 4).
The numbers of migrating neutrophils in zebrafish treated
with the positive control DEX, compound 8a and compound
8b (13, 16 and 12, respectively) were smaller than the control
group (25), indicating that compounds 8a, 8b and DEX had
comparable anti-inflammatory effects in EGFP transgenic
zebrafish.
in the model group at each time point, especially at 6, 8, and 10 h
(Fig. 5). Animals in the groups treated with compound 8b (low,
middle and high doses) also had noticeably less swelling than ani-
mals in the model group, but more than animals in the DEX group
at the same time points. Compared with the model group, animals
treated with compound 8b showed significant differences at 8 and
10 h. Compound 8b thus produced an anti-inflammatory effect at
all three doses and was comparable in efficacy to the positive con-
trol DEX.
In rat Freund ’s complete adjuvant model, the body weights of
animals in the model group, and in all three groups treated with
compound 8b, increased significantly, whereas the body weights
of animals in the positive control MTX group started to fall after
day 29. Body weights increased by 80.8, 70.8, and 58.2 g, respec-
tively, for animals treated with low, middle and high doses of
compound 8b. The increase of body weight in the low dose group
was comparable with that in the model group (80.8 vs 84.6 g) and
the increase of body weight in the high dose group was
In the rat carrageenan-induced paw edema model, animals
treated with the positive control DEX had less swelling than those
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Y. Jiang et al. / Bioorganic & Medicinal Chemistry Letters xxx (2018) xxx–xxx
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Fig. 2. Concentration–time curve of 8a (d), 7a (j), 8b (▲), and 7b (.) in HEPES buffer of pH 7.4.
Fig. 3. Concentration–time curve of 8a (d), 7a (j), 8b (▲), and 7b (.) in mouse plasma.
comparable with that in the MTX group (58.2 vs 59.2 g) (Fig. 6A).
In the model group, the degree of paw swelling was noticeably
increased. The degree of paw swelling was significantly decreased
after treatment with MTX or 8b for 4 days (P < .05, compared
with the model group). From day 10, the reduction of paw swel-
ling in animals treated with 8b (10 and 20 mg/kg) was compara-
ble with that in the MTX group, but the high dose of 8b (40 mg/
kg) was less effective. (Fig. 6B). Low dose 8b (10 mg/kg) thus
markedly relieved arthritis-induced inflammation, with minimal
effects on body weight.
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Y. Jiang et al. / Bioorganic & Medicinal Chemistry Letters xxx (2018) xxx–xxx
Fig. 5. Effect of compound 8b on the progress of acute inflammation induced by CG
in Sprague-Dawley rats. 30 rats were randomly divided into 5 groups, and orally
administered with positive control DEX (5 mg/kg) and compound 8b (10, 20 and 50
mg/kg) for 2 times at 2 h and 6 h after finishing injected carageen glue, respectively.
*P < .05, **P < .01, compared with the model group, SPSS 14.0 was used to analyze
the data. CG: Carrageen glue; DEX: dexamethasone.
Fig. 4. Effect of compounds 8a and 8b on the progress of traumatic inflammation in
EGFP transgenic zebrafish. 24 EGFP transgenic zebrafishes of 72 h development
period were divided into
4 groups of control group, positive control DEX,
compounds 8a and 8b. 2 ml positive control DEX (30 mg/mL), compounds 8a (5
mg/kg) for 5 weeks did not cause appreciable toxicity, as judged
by body weight (Fig. 7).
l
g/mL) and 8b (0.5 lg/mL) were added to cultured zebrafishes well for 3 h,
respectively. *P < .05, **P < .01, compared with the model group, SPSS 14.0 was used
to analyze the data. EGFP: enhanced green fluorescent protein; DEX:
dexamethasone.
Isothiocyanate prodrugs 8a and 8b were successfully synthe-
sized. Stability studies indicated that 8a and 8b were stable under
acidic solution but were gradually converted to the parent drugs at
higher pH. This suggests that the prodrugs should be stable in gas-
tric juice but should slowly release the active substances in blood.
Compounds 8a and 8b produced anti-inflammatory effects in the
neutrophilic inflammation model in EGFP transgenic zebrafishes.
Compound 8b showed comparable anti-inflammatory activity with
DEX, at a dose 300-fold lower than that of DEX. In the carrageenan-
induced paw edema model in rats, compound 8b showed anti-
inflammatory effects at low, medium and high doses, but displayed
more adverse effects than DEX. At the lowest dose (10 mg/kg), 8b
Toxicology of the compound 8b were evaluated in mice. Mice
treated for 5 weeks with compound 8b (200 mg/kg) survived,
and body weights in both the control group and the animals trea-
ted with compound 8b increased significantly. Body weight
increased more slowly in animals treated with compound 8b
group, but from day 30, body weights in the treatment with com-
pound 8b group were comparable with those in the control group.
Therefore, treatment with compound 8b at the tested dose (200
Fig. 6. Effect of compound 8b on the progress of Rheumatoid arthritis induced by FCA in Wistar rats. 30 Wistar rats were randomly divided into 5 groups, and orally
administered with positive control MTX (1 mg/kg) and compound 8b (10, 20 and 40 mg/kg) from day 16 after the initial inflammation, respectively. (A) The effect of
compound 8b on the increase of the body weight. (B) The effect of compound 8b on the paw swelling degree. the paw swelling degree was evaluated at two days intervals. *P
< .05, **P < .01, compared with the FCA group, SPSS 14.0 was used to analyze the data. FCA: Freund’s complete adjuvant; MTX: methotrexate.
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Y. Jiang et al. / Bioorganic & Medicinal Chemistry Letters xxx (2018) xxx–xxx
5
the Natural Science Foundation of Tianjin (No. 16JCQNJC13800 to
CG.B.).
A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at https://doi.org/10.1016/j.bmcl.2018.01.009.
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This work was supported by the National Natural Science Foun-
dation of China (NSFC) (No. 81573282 to Y.C.), the National Science
Fund for Distinguished Young Scholars (No. 81625021 to Y.C.) and
Please cite this article in press as: Jiang Y., et al. Bioorg. Med. Chem. Lett. (2018), https://doi.org/10.1016/j.bmcl.2018.01.009