Synthesis and antiproliferative activity of some novel triazole derivatives from dehydroabietic acid

Article abstract of DOI:10.3390/molecules19022523

Dehydroabietic acid (DHA) is a naturally occurring diterpene with different and relevant biological activities. Previous studies have shown that some DHA derivatives display antiproliferative activity. However, the reported compounds did not include triazole derivatives. Starting from DHA (8,11,13-abietatrien-18- oic acid), and its alcohol dehydroabietinol (8,11,13-abietatrien-18-ol), four alkyl esters were prepared. The alkyl terpenes were treated with different aromatic azides to synthesize hybrid compounds using click chemistry. Some 16 new DHA hybrids were thus synthesized and their structures were confirmed by spectroscopic and spectrometric means. The antiproliferative activity of the new compounds was assessed towards human cell lines, namely normal lung fibroblasts (MRC-5), gastric epithelial adenocarcinoma (AGS), lung cancer (SK-MES-1) and bladder carcinoma (J82) cells. Better antiproliferative effect was found for compound 5, with an IC₅₀ of 6.1 μM and selectivity on SK-MES-1 cells. Under the same experimental conditions, the IC₅₀ of etoposide, was 1.83 μM.

Full text of DOI:10.3390/molecules19022523

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Molecules 2014, 19, 2523-2535; doi:10.3390/molecules19022523
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molecules
ISSN 1420-3049
www.mdpi.com/journal/molecules
Article
Synthesis and Antiproliferative Activity of Some Novel Triazole
Derivatives from Dehydroabietic Acid
Mariano Walter Pertino 1,*, Valery Verdugo 1, Cristina Theoduloz 2 and
Guillermo Schmeda-Hirschmann 1
1
Instituto de Química de Recursos Naturales, Universidad de Talca, Casilla 747, Talca, Chile
2
Facultad de Ciencias de la Salud, Universidad de Talca, Casilla 747, Talca, Chile
* Author to whom correspondence should be addressed; E-Mail: mwalter@utalca.cl;
Tel.: +56-71-2241-8866.
Received: 24 January 2014; in revised form: 18 February 2014 / Accepted: 18 February 2014 /
Published: 21 February 2014
Abstract: Dehydroabietic acid (DHA) is a naturally occurring diterpene with different and
relevant biological activities. Previous studies have shown that some DHA derivatives
display antiproliferative activity. However, the reported compounds did not include triazole
derivatives. Starting from DHA (8,11,13-abietatrien-18-oic acid), and its alcohol
dehydroabietinol (8,11,13-abietatrien-18-ol), four alkyl esters were prepared. The alkyl
terpenes were treated with different aromatic azides to synthesize hybrid compounds using
click chemistry. Some 16 new DHA hybrids were thus synthesized and their structures
were confirmed by spectroscopic and spectrometric means. The antiproliferative activity of
the new compounds was assessed towards human cell lines, namely normal lung
fibroblasts (MRC-5), gastric epithelial adenocarcinoma (AGS), lung cancer (SK-MES-1)
and bladder carcinoma (J82) cells. Better antiproliferative effect was found for compound
5, with an IC50 of 6.1 μM and selectivity on SK-MES-1 cells. Under the same experimental
conditions, the IC50 of etoposide, was 1.83 µM.
Keywords: dehydroabietic acid; click chemistry; antiproliferative activity
1. Introduction
Synthesis or modification of known anticancer drugs is an important aspect of research. However,
to date a vast amount of synthetic work has contributed to relatively small improvements over the

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Molecules 2014, 19
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prototype drugs [1]. There is a continuous need for new templates to be used in the design of potential
chemotherapeutic agents. Dehydroabietic acid (DHA), a naturally occurring resin diterpene, and its
derivatives, exhibit a broad spectrum of biological effects, including antiulcer [2,3], antimicrobial [4],
antiviral [5], cytotoxic [3,6] and antitumor activities [7]. A recent patent on the use of abietic acid and
derivatives as antitumor agents has been published [8].
The search for drugs based on natural products often has the problem of involving slow and
complex synthetic pathways [9]. Click chemistry offers an alternative approach because it is a simple
and selective reaction between azides and alkynes to generate triazoles. The 1,4-disubstituted triazole
moiety is well known in medicinal chemistry. Before the discovery of the click chemistry reaction,
more than 7,000 compound bearing a 1,4-disubstituted 1H-1,2,3-triazole had been reported. The
triazole unit is stable to oxidation and reduction, and is quite resistant to metabolic degradation [10]. In
the design of novel prototypes for new antitumor drugs, triazole rings became increasingly relevant in
studies published in 2013 [11–13]. Stefely et al. [14] reported antiproliferative triazoles with an IC50 of
46 nM against MCF-7 human breast tumor cells. Moreover, the anti-angiogenic drug, 1,2,3-triazole
carboxyamidotriazole is being investigated in clinical trials as a potential anticancer drug [15,16].
Recently, thiourea α-aminophosphonate derivatives were attached to DHA, searching for antitumor
agents [7]. The authors measured cytotoxicity as the protocols for cell toxicity (cytotoxicity) and
antiproliferative effects are different. Moreover, antitumoral activity requires animal experiments. The
compounds showed moderate to high toxicity on NCI-H460, A549, HepG2 and SKOV3 cells. Some of
the compounds were more toxic than the commercial anticancer drug 5-fluorouracil.
The click chemistry approach to design new antiproliferative/anticancer agents has acquired
relevance in the last few years. In previous works, we reported the antiproliferative activity of dimeric
labdane diterpenes, some of them synthesized using click chemistry [17] and 1,2,3-triazole-substituted
oleanolic acid derivatives [18]. Kádár et al. [19] synthesized steroidal triazole derivatives to investigate
their antiproliferative effects. The most promising compounds were studied to evaluate its mechanism
of action, where apoptosis plays an important role. Moreover Li et al. [20] explored the addition of
triazole rings to naphthalimide to obtain hybrids with cytotoxic activity. One of the new hybrids was
more cytotoxic that the reference compound amonafide against the MCF-7, Hela and liver cancer
7721 cells, with IC50 values in the range of 0.323–1.02 µM. Under the same experimental conditions,
the IC50 of amonafide was in the range of 1.68–4.27 µM. The results points out the potential of click
chemistry to design new bioactive agents. However, other tools have to be included for a more rational
and effective drug design, including molecular modeling and receptor studies. The aim of this study
was to synthesize a series of hybrid molecules combining DHA with different aromatic-substituted
triazoles and to assess the antiproliferative effect of the new compounds in a panel of human tumor cell
lines. Recently, one of the alkyl esters prepared in this work (DHA propargyl ester) was incorporated
to caprolactone polymer (polycaprolactone, PCL) by click chemistry reaction. The integration of DHA
to PLC increases the glass-transition temperature of PCL and does excellent hydrophobicity and good
degradability to the polymer [21].

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2. Results and Discussion
Starting from DHA and its alcohol, four alkyl esters were prepared and then treated with different
aromatic azides using click chemistry to produce 16 hybrid abietane-triazole compounds (Schemes 1
and 2). The new compounds were obtained in moderate to good yields. The method proved to be
suitable to obtain series of derivatives from diterpenes and can be applied to other natural products to
increase structural diversity. Compounds 1–16 are described for the first time. The structures of all the
compounds were confirmed by spectroscopic and spectrometric means.
Scheme 1. Preparation of DHA derivatives 1–4.
a
b
OH
O
R
O
N
N
O
O
O
n
n
N
1
2
3
4
n:1 R: Phenyl
n:2 R: Phenyl
n:1
n:2
R: Methyl phenyl sulfide
R: Methyl phenyl sulfide
Reagents and conditions: (a) appropriate alkyne alcohol, DCC, DMAP, CH2Cl2, 65%–77%;
(b) appropriate aromatic azide, CuSO4·5H2O, sodium ascorbate, t-BuOH/H2O 1:1, 59%–70%.
Scheme 2. Preparation of dehydroabietinol derivatives 5–16.
b
c
R
N
O
O
O
O
N
OH
N
n
n
a
5
6
7
8
9
n:1 R: Phenyl
n:3 R: Phenyl
n:1 R: Benzyl
n:3 R: Benzyl
n:1 R: Methyl phenyl sulfide
10 n:3 R: Methyl phenyl sulfide
11 n:1 R: p-anisidyl
12 n:3 R: p-anisidyl
13 n:1 R: m-chlorophenyl
14 n:3 R: m-chlorophenyl
15 n:1 R: p-toluenesulfonyl
16 n:3 R: p-toluenesulfonyl
OH
ADH
O
Reagents and conditions: (a) LiAlH4, THF, 80 °C, 86%; (b) appropriate alkyne acid, DCC, DMAP,
CH2Cl2, 60%–81%; (c) appropriate aromatic azide, CuSO4·5H2O, sodium ascorbate, t-BuOH/H2O
1:1, 54%–89%.

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The compounds were then assessed for antiproliferative activity towards the following human cell
lines: normal lung fibroblasts (MRC-5), gastric epithelial adenocarcinoma (AGS), lung cancer
(SK-MES-1) and bladder carcinoma (J82) cells. IC50 values > 100 µM were considered inactive (Table 1).
Table 1. Antiproliferative activity of dehydroabietic acid derivatives 1–16 against MRC-5
normal fibroblasts and selected tumor cell lines. a
(IC50 ± SD, µM) b
Compound
MRC-5
85.8 ± 4.6
>100
AGS
79.9 ± 4.8
90.2 ± 5.4
94.4 ± 5.4
66.0 ± 4.4
44.5 ± 1.9
>100
SK-MES-1
77.3 ± 4.7
>100
J82
46.3 ± 2.3
76.3 ± 5.9
>100
1
2
3
>100
>100
4
50.2 ± 3.1
17.1 ± 0.6
>100
28.9 ± 2.1
6.1 ± 0.3
>100
28.1 ± 1.7
83.3 ± 3.3
>100
5
6
7
>100
36.7 ± 1.5
35.9 ± 1.7
47.7 ± 2.9
58.9 ± 3.4
>100
39.7 ± 2.7
37.0 ± 2.6
43.6 ± 2.9
42.7 ± 2.9
>100
25.6 ± 1.9
26.8 ± 1.8
49.9 ± 2.9
33.9 ± 1.7
>100
8
>100
9
73.3 ± 3.7
>100
10
11
>100
12
>100
>100
>100
>100
13
>100
>100
>100
>100
14
15
>100
>100
>100
>100
82.9 ± 4.5
68.8 ± 4.5
0.33 ± 0.02
74.9 ± 3.7
56.9 ± 3.3
0.58 ± 0.02
84.4 ± 5.1
74.2 ± 5.4
1.83 ± 0.09
>100
16
75.0 ± 6.1
3.49 ± 0.16
Etoposide c
a
Cell lines: normal lung fibroblasts (MRC-5), gastric epithelial adenocarcinoma (AGS), lung cancer
b
(SK-MES-1) and bladder carcinoma (J82) cells; Results are expressed as mean values ± SD. Each
concentration was tested in sextuplicate together with the control and repeated two times in separate
experiments; c Reference compound.
Among the triazoles prepared with the diterpene acid, the derivatives 1 and 4 presented
antiproliferative activity towards all of the selected cell lines. The effect, however, is low. Compounds
2 and 3, differing in the number of CH2 groups of the linker or in the aromatic moiety were inactive
against MRC-5 and SK-MES1cells. When comparing the pairs 1–2 and 3–4 bearing one or two CH2
units as linkers, the effect of 1 was higher than that of 2, suggesting that a longer chain decreases
activity when R is a phenyl group. However, an opposite effect was found for the methyl phenyl
sulfide derivatives. The best compound of this group was the derivative 4, with a (CH2)2 linker and a
methyl phenyl sulfide substituent in the triazole ring. More derivatives with longer linker chains need
to be prepared to have a clearer picture of the effect of this structural feature on the antiproliferative effect.
The compounds 5–16, were prepared with the diterpene alcohol. Among them, the compounds 11–14
were inactive. All of them contain electron donors such as a methoxy group or a chlorine atom in the
aromatic ring. Compounds 7 and 8 showed similar activity against AGS, SK-MES-1 and J82 cells and
were inactive against fibroblasts, showing a good selectivity. This fact suggests that the length of the
CH2 linker from 1 to 3 units do not have a relevant influence in the antiproliferative activity. The same