Synthesis and biological evaluation of novel triazole derivatives as antifungal agents

Article abstract of DOI:10.1016/j.cclet.2013.01.015

A series of 1-(benzylamino)-2-(2,4-difluorophenyl)-3-(1H-1,2,4-triazol-1- yl)propan-2-ols compounds were synthesized and evaluated for their antifungal activities in vitro. The results showed that compounds 6A and 6B exhibited good antifungal activity. Compound 6A8 showed the strongest antifungal activity, which was significantly higher than that of the lead compounds and positive-control drugs Fluconazole and Itraconazole. In particular, the antifungal activity of compound 6A8 against Candida albicans and Candida krusei (MIC80 both at 0.00097 μg/mL) was 515 and 64 times that of Fluconazole, respectively. The structure-activity relationships of the synthesized compounds were discussed, and the docking model of the target compounds with fungal lanosterol 14α-demethylase (CYP51) was analyzed.

Full text of DOI:10.1016/j.cclet.2013.01.015

Chinese Chemical Letters 24 (2013) 219–222
Contents lists available at SciVerse ScienceDirect
Chinese Chemical Letters
journal homepage: www.elsevier.com/locate/cclet
Original article
Synthesis and biological evaluation of novel triazole derivatives as antifungal
agents
a
a
a
a
a
a,
Hui Tang b,1, Can-Hui Zheng a,1, , Xiao-Hui Ren , Jia Liu , Na Liu , Jia-Guo Lv , Ju Zhu , You-Jun Zhou
*
*
a School of Pharmacy, Second Military Medical University, Shanghai 200433, China
b Pharmacy, Provincial Hospital Affiliated to Shandong University, Jinan 250021, China
A R T I C L E I N F O
A B S T R A C T
Article history:
A series of 1-(benzylamino)-2-(2,4-difluorophenyl)-3-(1H-1,2,4-triazol-1-yl)propan-2-ols compounds
were synthesized and evaluated for their antifungal activities in vitro. The results showed that
compounds 6A and 6B exhibited good antifungal activity. Compound 6A8 showed the strongest
antifungal activity, which was significantly higher than that of the lead compounds and positive-control
drugs Fluconazole and Itraconazole. In particular, the antifungal activity of compound 6A8 against
Received 26 October 2012
Received in revised form 2 December 2012
Accepted 17 December 2012
Keywords:
Candida albicans and Candida krusei (MIC80 both at 0.00097
mg/mL) was 515 and 64 times that of
Triazole
Fluconazole, respectively. The structure–activity relationships of the synthesized compounds were
Antifungal
discussed, and the docking model of the target compounds with fungal lanosterol 14
(CYP51) was analyzed.
a-demethylase
Structure–activity relationship
CYP51
ß 2013 Can-Hui Zheng. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights
reserved.
1. Introduction
the active cavity of the target enzyme, whereas 2,4-difluorophenyl
binds to the hydrophobic pocket. The configuration of the
During the past two decades, the incidence and death rate of
deep fungal infection have drastically increased, which has become
a main cause of death in major diseases such as AIDS and
tumors [1,2]. Such alarming data demonstrates the importance of
antifungal drugs in clinical anti-infective treatment. Currently,
azole antifungal drugs are widely used in clinical treatment [3]; in
particular, Fluconazole, Itraconazole, Voriconazole, etc. (Fig. 1), are
now preferred as first-line antifungal therapy [4,5]. Recently, new
azole drugs such as Posaconazole and others have also entered the
market [6]. However, the extensive use of azole antifungal drugs
has led to the resistance of clinical pathogenic fungi to drug
treatment, one of the main causes of clinical treatment failure [4,7].
Therefore, development of new azole drugs for clinical treatment is
a pressing need.
remaining side chain varies considerably and it binds to the entry
pathway of the target enzyme and can largely affect its ability to
bind target enzyme [9].
Previous studies showed that 1-(benzyl(prop-2-ynyl)amino)-2-
(2,4-difluorophenyl)-3-(1H-1,2,4-triazol-1-yl)propan-2-ol com-
pounds (Fig. 2) exhibited good antifungal activity [10]. In this
study, we designed and synthesized a series of derivatives of these
compounds (Fig. 2) by replacing the aromatic ring on the side
chain, changing the distance between the aromatic ring and the
nitrogen atom and replacing different groups on the nitrogen. The
antifungal activity of target compounds and their preliminary
structure–activity relationships were investigated, and the dock-
ing model of the target compounds with CYP51 was analyzed.
Azole antifungal drugs block the biosynthesis of fungal cell
membranes and suppress fungal growth by inhibiting CYP51
which is a critical enzyme in the ergosterol synthesis pathway [8].
Researches indicated that the triazole ring and 2,4-difluorophenyl
group were the pharmacophores of antifungal agents. The 1,2,4-
triazole ring binds to the Fe atom of the heme prosthetic group in
2. Experimental
The synthesis of the target compounds was performed as shown
in Scheme 1. Key intermediate 4 was synthesized via a known
route [11]. Intermediate 4 was reacted with different alkylamines
in the presence of triethylamine, then treated with concentrated
HCl to provide the key intermediates 5 [10]. Lastly, in the presence
of KI and K2CO3, intermediate 5 reacted with different substituted
benzyl chlorides, and target compound 6 was obtained. Com-
pounds 6A1-6A4 are known lead compounds and served as
positive controls; the remaining 31 compounds were all new.
* Corresponding authors.
E-mail addresses: canhui_zheng@yahoo.com.cn (C.-H. Zheng),
zhouyoujun2006@yahoo.com.cn (Y.-J. Zhou).
1
These authors contributed equally to this work.
1001-8417/$ – see front matter ß 2013 Can-Hui Zheng. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
http://dx.doi.org/10.1016/j.cclet.2013.01.015

220
H. Tang et al. / Chinese Chemical Letters 24 (2013) 219–222
Fig. 1. Triazole antifungal drugs.
Fig. 2. The target compounds.
Supplementary data associates with this article can be found in
supporting information.
Compound 6A8 exhibited significantly higher antifungal activity
against most of the strains (including A. fumigatus), than did the
positive-control drugs Fluconazole and Itraconazole. In particular,
the antifungal activity of compound 6A8 against Candida albicans
and C. krusei was respectively 515 and 64 times that of Fluconazole
and was several times stronger than that of lead compounds 6A1–
6A4 as well.
As most 6A compounds exhibited good in vitro antifungal
activity, this supported our contention that replacing the side
chain on the nitrogen with propargyl could indeed substantially
increase the activity of the compound [10]. However, the
antifungal activity of most 6B compounds, although higher than
Fluconazole, was noticeably lower than 6A compounds. This
indicated that replacing the side chain on nitrogen with
cyanomethyl was worse for antifungal activity than replacing
with propargyl. On the other hand, replacing the side chain on
nitrogen with a long hydrocarbon chain almost eliminated the
antifungal activity of the compound, which was demonstrated by
3. Results and discussion
All the new target compounds were tested in vitro against seven
common human pathogenic fungi [12,13], and the MIC80 values are
shown in comparison with that of Fluconazole and Itraconazole in
Table 1.
Compounds 6A1–6A4 were known lead compounds that
showed good activity [10]. They were tested for antifungal activity
for comparison purposes in our study. As shown in Table 1, the new
compounds, 6A7–6A8, 6A12–6A14, 6B1–6B9, 6B11–6B13 showed
high-efficiency, broad-spectrum antifungal activity against six
fungal strains (except Aspergillus fumigatus), compounds 6A5–6A6,
6A9–6A11 and 6B10 showed middle-level antifungal activity and
compounds 6C, 6D, 6E showed low antifungal activity. Noticeably,
the activity of compounds 6A7–6A8 was especially strong.
Scheme 1. The synthesis route of the target compounds. Reagents and conditions: (a) ClCH2COCl, AlCl3, 50 8C, 5 h, yield 85%; (b) 1H-1,2,4-triazole, K2CO3, C6H5CH3, reflux, 5 h,
yield 82%; (c) (CH3)3SOI, NaHCO3, C6H5CH3, 60 8C, 3 h; (d) CH3SO3H, 0 8C, 1 h, yield 75%; (e) alkylamine, Et3N, CH3CH2OH, reflux, 5-6 h; (f) HCl (g), yield 80-90%; (g) substituted
benzyl chloride, KI, K2CO3, CH3CN, reflux, 5-6 h, yield 30-50%.

H. Tang et al. / Chinese Chemical Letters 24 (2013) 219–222
221
Table 1
The structure and in vitro antifungal activity (MIC80
,
mg/mL) of target compounds.
No.
Compound structure
Antifungal activity (MIC80, mg/mL)
R1
n
1
X
C
R2
H
C. alb. SC5314
C. alb. Y0109
C. neo.
C. tro.
T. rub.
C. par.
C. kru.
A. fum.
6A1
0.0039
0.0039
0.25
0.0625
1
0.0625
0.0039
64
>64
64
6A2
1
1
1
1
1
1
1
1
1
1
2
2
2
1
C
C
C
C
C
C
C
C
C
N
C
C
C
C
4-F
3-F
2-F
0.0156
0.0039
0.0156
0.0156
0.25
0.0156
0.0156
0.0625
4
0.0625
0.0625
0.0156
0.25
0.25
64
64
16
16
16
16
1
0.0625
0.0625
0.0625
16
0.0156
0.0156
0.0156
0.0156
0.25
6A3
0.0625
6A4
0.0625
>64
>64
>64
64
6A5
4-C(CH3)3
4-CF3
3,4-2F
2,4-2F
3,4-2Cl
2,3-2Cl
H
1
4
6A6
0.25
1
1
6A7
0.0039
0.00097
0.25
0.0156
0.00097
0.25
0.0625
0.0156
1
0.25
0.25
4
0.0625
0.0156
1
0.0156
0.0039
0.25
6A8
16
6A9
16
16
64
64
>64
16
64
>64
>64
>64
>64
>64
64
6A10
6A11
6A12
6A13
6A14
6B1
0.25
0.25
0.0625
0.0625
0.0625
0.0625
0.0156
0.0625
16
1
0.25
0.25
1
1
0.25
0.25
0.0156
0.0625
0.0625
0.0156
0.25
4-F
0.0625
0.0625
0.0625
0.25
0.0625
0.0625
0.0625
0.25
4
3-F
1
0.25
0.25
0.25
2-F
0.25
1
H
>64
N
N
N
N
N
N
N
N
N
N
N
N
N
6B2
1
1
1
1
1
1
1
1
1
2
2
2
C
C
C
C
C
C
C
C
N
C
C
C
4-F
0.0625
0.0156
0.0625
0.0156
0.0625
0.0625
0.0625
0.0625
0.25
0.0625
0.0625
0.0625
0.0156
0.0625
0.0625
0.0625
0.0625
0.25
0.0625
0.0625
0.0156
0.0625
0.0625
0.25
0.25
0.25
1
>64
16
0.25
0.0625
0.0156
0.0625
0.0156
0.0625
0.0625
0.0625
0.0625
0.0625
0.0156
0.0625
0.0625
>64
>64
>64
64
6B3
3-F
0.0625
0.25
6B4
2-F
64
6B5
4-Cl
4-OCH3
4-CH3
3,4-2F
2,4-2F
H
0.25
4
16
0.0625
0.25
6B6
16
>64
>64
>64
64
6B7
1
>64
>64
4
0.25
6B8
0.25
0.25
0.25
4
0.25
6B9
0.0625
0.25
0.0625
4
6B10
6B11
6B12
6B13
64
>64
64
4-F
0.0156
0.0625
0.0625
0.0156
0.0625
0.0156
0.0625
0.0625
0.0625
0.25
4
0.0156
0.0625
64
0.0625
0.25
3-F
>64
>64
2-F
0.25
0.25
6C1
C6H13
C6H13
C7H15
C7H15
C8H17
C8H17
/
1
1
1
1
1
1
C
C
C
C
C
C
/
4-F
4-Cl
4-F
4-Cl
4-F
4-Cl
/
16
4
/
64
64
64
64
64
64
64
16
1
>64
>64
>64
>64
>64
>64
>64
4
6C2
4
4
/
4
16
1
6D1
4
4
/
4
16
1
6D2
1
1
/
16
4
1
6E1
4
4
/
64
64
1
6E2
1
1
/
4
4
1
Fluconazole
Itraconazole
/
0.5
0.5
0.25
0.0625
1
0.25
0.25
0.25
0.0156
0.25
0.0625
/
/
/
/
0.0625
0.0625
0.0625
C. alb., SC5314 and Y0109, Candida albicans; C. neo., Cryptococcus neoformans; C. tro., Candida tropicalis; T. rub., Trichophyton rubrum; C. par., Candida parapsilosis; C. kru., Candida
krusei; A. fum., Aspergillus fumigatus.
the low antifungal activity of compounds 6C, 6D and 6E.
Combining the results from this study and those from previous
research [10,14,15], we proposed that the optimal strategy to
maximize the antifungal activity of compound was to replace the
side chain on nitrogen with a group of three carbon atoms.
The substituted group on the side chain benzene ring also
contributed significantly to the compound activity. Compounds
6A7–6A8 showed highest activity, indicating that the
difluoro-substituted group was especially favorable for antifun-
gal activity. In addition, based on the activity comparisons
between 6A11 and 6A1, 6B10 and 6B1, we obtained
a
preliminary conclusion that to replace the side-chain benzene
ring with a heteroaromatic ring was unfavorable for antifungal
activity. Based on the activity comparisons between 6A12–6A14
and 6A2–6A4, and 6B11–6B13 and 6B2–6B4, we found that to
increase the distance between the side-chain benzene ring and

222
H. Tang et al. / Chinese Chemical Letters 24 (2013) 219–222
compound. Compound 6A8 provides a good starting point for
further structural optimization.
Acknowledgments
This work was supported by the National Natural Science
Foundation of China (Nos. 20972187 and 30901859), Shanghai
Natural Science Foundation (No. 09ZR1438800) and Shandong
Natural Science Foundation (No. ZR2012HQ026).
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.cclet.2013.01.015.
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(MIC80 = 0.00097
mg/mL) which was significantly stronger than
that of the lead compounds and positive-control drugs. The side
chain on nitrogen with propargyl of compounds is important for
their antifungal activity. Replacing the side chain on nitrogen with
cyanomethyl was worse for antifungal activity than replacing with
propargyl and replacing the side chain on nitrogen with a long
hydrocarbon chain almost eliminated the antifungal activity of the