Click chemistry-based synthesis and anticancer activity evaluation of novel C-14 1,2,3-triazole dehydroabietic acid hybrids

Article abstract of DOI:10.1016/j.ejmech.2017.07.049

A concise and efficient synthetic approach has been established to readily access a series of novel C-14 1,2,3-triazole-tethered dehydroabietic acid derivatives in moderate to high yields. In vitro antiproliferative activity evaluation indicated that most of the hybrids exhibited potent inhibitory activities in a variety of cancer cell lines with low micromolar to submicromolar IC₅₀ values. Further studies demonstrated that some of these analogues such as 20, 21, and 24 were also effective against adriamycin-resistant MCF-7 clone at low concentrations in a dose-dependent manner. Notably, the most potent compound 24, which possesses a 3-(tert-butoxycarbonylamino)phenyl-substituted triazole moiety, not only exhibited obviously improved IC₅₀ values ranging from 0.7 to 1.2 μM against a panel of tested cancer cells, but also showed very weak cytotoxicity on normal cells. Preliminary mechanism studies indicated that compound 24 could induce apoptosis in MDA-MB-231 cells and was worth developing into a novel natural product-like anticancer lead by proper structure modification.

Full text of DOI:10.1016/j.ejmech.2017.07.049

Accepted Manuscript
Click chemistry-based synthesis and anticancer activity evaluation of novel C-14
1,2,3-triazole dehydroabietic acid hybrids
Wei Hou, Zhi Luo, Guanjun Zhang, Danhui Cao, Di Li, Haoqiang Ruan, Benfang
Helen Ruan, Lin Su, Hongtao Xu
PII:
S0223-5234(17)30571-8
10.1016/j.ejmech.2017.07.049
EJMECH 9612
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 11 April 2017
Revised Date: 20 July 2017
Accepted Date: 22 July 2017
Please cite this article as: W. Hou, Z. Luo, G. Zhang, D. Cao, D. Li, H. Ruan, B.H. Ruan, L. Su,
H. Xu, Click chemistry-based synthesis and anticancer activity evaluation of novel C-14 1,2,3-
triazole dehydroabietic acid hybrids, European Journal of Medicinal Chemistry (2017), doi: 10.1016/
j.ejmech.2017.07.049.
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ACCEPTED MANUSCRIPT

ACCEPTED MANUSCRIPT
Click chemistry-based synthesis and anticancer activity
evaluation of novel C-14 1,2,3-triazole dehydroabietic acid
hybrids
Wei Houa, Zhi Luob, Guanjun Zhangc, Danhui Caoa, Di Lia, Haoqiang Ruana, Benfang Helen Ruan a, Lin Sua,*,
Hongtao Xu d,e
*
aCollege of Pharmaceutical Science, Collaborative Innovation Center of Yangtza River Delta Region Green Pharmaceuticals, Zhejiang
University of Technology, Hangzhou 310014, PR China
bShanghai Evergene Biotech Co,. Ltd., Shanghai, 201499, PR China
cCollege of Chemical Engineering and Materials Science, Tianjin University of Science & Technology, Tianjin, 300457, PR China
dShanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, Shanghai 201210, PR China
eState key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, PR
China
*Corresponding author
E-mail: xuhongtao2010@126.com, htxu2010@hotmail.com (H.T. Xu), sulin8023@zjut.edu.cn (L. Su)
Abstract
A concise and efficient synthetic approach has been established to readily access a series of
novel C-14 1,2,3-triazole-tethered dehydroabietic acid derivatives in moderate to high yields. In
vitro antiproliferative activity evaluation indicated that most of the hybrids exhibited potent
inhibitory activities in a variety of cancer cell lines with low micromolar to submicromolar IC50
values. Further studies demonstrated that some of these analogues such as 20, 21, and 24 were
also effective against adriamycin-resistant MCF-7 clone at low concentrations in a dose-dependent
manner. Notably, the most potent compound 24, which possesses
a
3-(tert-
butoxycarbonylamino)phenyl-substituted triazole moiety, not only exhibited obviously improved
IC50 values ranging from 0.7 to 1.2 µM against a panel of tested cancer cells, but also showed very
weak cytotoxicity on normal cells. Preliminary mechanism studies indicated that compound 24
could induce apoptosis in MDA-MB-231 cells and was worth developing into a novel natural
product-like anticancer lead by proper structure modification.
Key Words: terpenoids, dehydroabietic acid, cancer, 1,2,3-triazole, synthesis, click
chemistry, drug resistance
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1. Introduction
Cancer is characterized by its high incidence and mortality [1]. Despite the availability of
various chemotherapeutic drugs, the rapid development of multidrug resistance (MDR) and
associated acute side effects of clinical used anticancer drugs (including the recently introduced
drugs) are still the major obstacles to effective cancer chemotherapy [2]. Consequently, the
discovery and development of new effective anticancer agents and/or therapies to address the
unmet demands is always the major goal of medicinal chemists. On the other hand, natural
products (NPs) are structural-complex molecules that have a profound impact upon medicinal
chemistry. Their diversity three-dimensional shape, functionality, stereochemistry, as well as
various interesting biological activities have always provided medicinal chemists a reliable source
of inspiration in their search for new drug-like molecules [3]. It is reported that approximately 60%
of clinically used antitumor drugs originated from NPs. For example, vincristine, vinblastine,
irinotecan, etoposide, doxorubicin and paclitaxel are well-known naturally derived
chemotherapeutic agents in clinical use [4].
Terpenoids are the most abundant and widely distributed naturally occurring interesting
metabolites. As the largest class of NPs, it contains approximately 25,000 chemical structures,
thus exhibiting extensive pharmaceutical activities [5]. Among them, abietic acid (AA, 1) and
dehydroabietic acid (DHAA, 2) are natural resin acids which can be easily obtained from pinus
rosin or disproportionated rosin (Figure 1). It is reported that AA, DHAA and their derivatives
showed a broad spectrum of bioactivities such as anti-inflammatory, antiviral, antimicrobial, BK
channel-opening and especially antitumor activities [6]. As a result, AA and DHAA have been
widely used as the starting material for the construction of several NPs and structurally diverse
natural product-like (NPL) molecules library [7].
Figure 1. Structures of abietic acid (AA) (1) and dehydroabietic acid (DHAA) (2)
In general, the search for new drugs based on structural modification of NPs has often been
hindered by the tedious and complicated synthetic pathways [8]. “Click chemistry”, a concept
initiated by Barry K. Sharpless [9], provided a nearly perfect strategy to perform selective
modification on structural complex NPs under mild conditions, even in the presence of reactive
functionalities. Of particular interest is the copper(I)-catalyzed alkyne-azide [3+2] cycloaddition
(CuAAC) reaction, which provides 1,4-disubstituted 1,2,3-triazoles [10]. This well-known
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pharmacophore is quite resistant to metabolic degradation and capable of participating in dipole-
dipole interactions as well as hydrogen bonding, thus providing additional advantages including
cell permeability improvement and target binding [11].
Considering the above benefits and in continuation of our interest in searching for the
anticancer pharmacological effects of abietane diterpenoids derivatives [12], we therefore
conducted a modification via integrating dehydroabietic acid (DHAA) and triazole pharmacophore
into one molecule to generate a novel C-14 1,2,3-triazole dehydroabietic acid hybrid scaffold
through CuAAC for further biological evaluation [13]. Herein, we described the synthesis of these
hybrids and the evaluation of their in vitro antiproliferative activities, selectivities over normal
human cells, potential against drug-resistance tumor cells, in vitro metabolic stabilities and
preliminary elucidation of their underlying mechanisms of cytotoxicity.
2. Results and discussion
2.1 Chemistry
The synthetic route towards the target DHAA-1,2,3-triazole hybrids was depicted in Scheme 1.
Briefly, the key intermediate C-14 azide acetamino-DHAA 8 was synthesized from DHAA by a
five-step streamlined manipulation. First, esterification of 2 provided the corresponding ester 3 in
98% yield. Subsequent nitration of ester 3 afforded a 3:2 mixture of 4 and 5. Reduction of 5
followed by amidation of the resulting C14-amine with chloracetyl chloride delivered 7, which
was treated with sodium azide to give the key intermediated 8 in high yield. Finally, reaction of 8
with various selected aliphatic and aromatic alkynes in the presence of CuSO4·5H2O and sodium
ascorbate produced target derivatives 9-34 in moderate to high yields. Further, preliminary drug-
likeness properties of the newly synthesized molecules were determined by in silico
Molinspiration online property calculation software and the data obtained were presented in Table
1 [14]. For most compounds, the parameters like hydrogen bond acceptor (n-ON), hydrogen bond
donor (n-OHNH), rotatable bonds (n-rotb) were in the range of Lipinski's rule of 5 [15]. As
expected, compounds with trimethyl chlorosilane- (13), benzyl- (14) and substituted phenyl- on
the C-4 position of the 1,2,3-triazole moiety showed higher lipophilicity (LogP 4.39-6.99), while
compounds with saturated alkyl- (10 and 11), ester- (12), pyridyl- (30, 31 and 32) and thienyl-
possessed lower lipophilicity (LogP 3.67-5.12). Morever, the polar surface area (TPSA) of all the
compounds (9-34) were less than 140 A2 [16].
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Scheme 1. Synthesis of 9-34. Reagents and conditions: (a) (MeO)2SO2, K2CO3, Acetone, rt., 12 h, 98%; (b) HNO3,
Ac2O, 0 °C to rt., 1 h, 4 (39%), 5 (20%); (c) Fe/NH4Cl, EtOH, reflux, 2 h, 75%; (d) chloracetyl chlride, THF, 0 °C
to rt., 2.5 h, 81%; (e) NaN3, DMF, 80 °C, 12 h, 77%; (f and g) alkyne, sodium ascorbate, CuSO4·5H2O, t-BuOH-
H2O (1:1), 40 °C.
2.2. Biological assays
From the synthetic route mentioned above, we obtained a series of novel DHAA-1,2,3-triazole
hybrids. To examine whether the substituents affected their cytotoxic activities, all the synthesized
hybrids were evaluated for their potential against four different human tumor cell lines including
ovary (SKOV-3), prostate (PC-3) and breast (MDA-MB-231 and MCF-7) by using MTT assay. 5-
Fu and triptonide were used as positive controls for comparison [17].
As expected, most of the hybrids exhibited moderate to high cytotoxic activities against four
tumor cell lines with IC50 values below 10 µM (Table 1). Among them, hybrids 20, 21 and 24
displayed significant cytotoxic activities against all the tested cancer cells with IC50 values ranging
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from 0.7±0.1 to 1.9±0.2 µM. Notably, compound 24 showed the strongest activity against MDA-
MB-231 cells with an IC50 of 0.7 ± 0.1 µM, indicating a higher potency compared with the
commercial anticancer drug 5-Fu. Meanwhile, the cytotoxic activities of the intermediates 6, 7 and
8 were also examined. The results revealed that 6 and 7 only displayed moderate cytotoxic
activities, suggesting that the introduction of 1,2,3-triazole moiety was beneficial to cytotoxicity.
Further, the effect of various substituents on the C-4 position of the 1,2,3-triazole moiety was
also examined and the structure-activity relationship (SAR) studies revealed that the introduction
of aromatic substituents was crucial for the potent cytotoxicity. Generally, the introduction of
electron-rich aromatic ring system could significantly increase the cytotoxicity. In particular,
compounds 19, 20, 21 and 24 showed promising IC50 values ranging from 1.0 to 2.3, 0.8 to 1.4,
1.4 to 1.9 and 0.7-1.2 µM respectively. Whereas, the introduction of electron-poor aromatic ring
system would hamper the cytotoxicity. For example, pyridyl- (30, 31 and 32) and nitrobenzyl-
substituted analogues (25 and 26) only showed weak or loss of cytotoxicities. In comparison, the
saturated alkyl- (10, 11, and 33), ester- (12) and trimethyl chlorosilane- (13) on the C-4 position of
the 1,2,3-triazole moiety were only associated with moderate increase in the growth inhibitory
effect.
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Table 1. IC50 and in silico pharmacokinetic evaluation of compounds 6-34.
IC50 (Mean ± S.D., µM)a
Compd
LogPb TPSAc
SKOV-3
6.1±1.4
69.7±4.3
>100
PC-3
25.4±0.9
71.2±0.9
>100
MDA-MB-231
24.1±3.3
>100
MCF7
31.1±1.2
51.6±5.5
52.11±8
15.6±2.8
>100
-----
-----
-----
3.67
4.47
4.11
3.97
6.04
5.72
5.20
5.36
5.34
5.31
5.21
5.23
5.28
6.17
4.39
6.07
5.13
5.16
5.24
6.38
6.99
4.05
4.13
3.91
5.12
4.98
-----
-----
-----
-----
6
7
60.7±4
-----
8
25.3±2.8
>100
49.0±1
>100
8.2±0.5
60.0±2.9
16.2±0.4
21.3±1.1
>100
138.73
86.12
86.12
112.43
86.12
86.12
86.12
86.12
86.12
86.12
95.36
95.36
95.36
95.36
115.22
124.45
131.94
131.94
104.59
86.12
86.12
99.01
99.01
99.01
136.82
86.12
-----
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
5-Fu
19.4±1.1
22.9±3.4
65.2±1.2
59.4±3.7
4.8±0.9
15.2±3.7
91.2±8.1
1.8±0.5
1.4±0.8
0.9±0.3
1.4±0.1
20.3±2.5
3.9±1.1
1.2±0.1
>100
12.3±4.0
15.4±2.1
94.3±7.9
>100
24.2±3.3
16.1±2.2
>100
31.9±6.4
3.0±0.6
10.0±2.6
>100
>100
3.4±0.6
8.7±1.3
>100
3.7±1
19.4±1.0
68.8±9.3
1.9±0.8
1.0±0.1
0.8±0.4
1.4±0.4
9.2±0.7
1.5±0.9
0.8±0.2
91.1±9.6
>100
2.6±0.6
2.3±0.3
1.1±0.5
1.9±0.2
12.1±1.3
10.1±2.0
0.9±0.1
>100
1.0±0.1
1.8±0.2
1.4±0.4
1.8±0.2
23.4±0.7
5.9±0.9
0.7±0.1
87.5±6.6
>100
77.3±5.7
3.3±0.1
45.3±0.5
16.0±7.5
>100
>100
5.2±0.9
9.1±0.4
11.1±1.0
>100
1.4±0.1
27.0±2.1
8.9±1.0
>100
8.9±2.0
12.1±3.3
9.5±2.0
>100
>100
30.0±5.5
>100
41.9±2.2
>100
>100
>100
>100
92.1±1.2
>100
>100
>100
>100
>100
57.1±1.0
24.5±1.8
0.03±0.004
>100
20.8±1.1
13.7±2.4
5.2±3.0
0.03±0.002
Triptonide 0.008±0.002 0.02±0.003
-----
aThe drug concentration required for 50% inhibition of cell viability. SKOV-3, PC-3, MDA-MB-231and MCF7
cells were treated with all the tested compounds for 72 h. Each experiment was run for at least three times
independently with nine concentrations, and the results are presented as mean ± S.D..
bLogP: Logarithm of partition coefficient of compound between n-octanol and water.
cTPSA: Topological polar surface area.
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On the basis of the results above, we further evaluated the in vitro antiproliferative activity of
the most potent compound 24 against another six tumor cell lines, which derived from different
human tissues, including glioma (U251), breast (SKBR-3), lung (A549), colon (HCT-116),
hepatoma (SMCC-7721) and prostate (LNCaP). The result showed that compound 24 strongly
inhibited the growth of all the tested cancer cells, exhibiting IC50 values ranging from 0.6 to 1.8
µM (Table 2). Meanwhile, the IC50 values in normal human foreskin fibroblast (HFF) and human
liver (HL-7702) cells were 5.4 µM and 7.8 µM respectively, implying there was a therapeutic
window to use compound 24.
Table 2. IC50 Values of compound 24 against several tumor cells and normal cells.
a
Cells
U251
SKBR-3
A549
LNCaP
SMCC-7721
HCT-116
HL-7702
HFF
IC50
SIb
4.5
8.4
6.1
12.4
7.6
7.9
----
----
SIc
3.1
5.8
4.2
8.6
5.2
5.4
----
----
1.8±0.2
0.9±0.1
1.3±0.3
0.6±0.3
1.1±0.2
1.0±0.2
7.8±1.3
5.4±0.8
aThe drug concentration required for 50% inhibition of cell viability, each experiment was run for at least three times independently with
nine concentrations, and the results are presented as mean ± S.D.
bSI: Selective index (IC50 on normal cells (HL-7702)/IC50 on tumor cells)
cSI: Selective index (IC50 on normal cells (HFF)/IC50 on tumor cells)
In principle, drug resistance to chemotherapy remains to be a major impediment to the
successful treatment of cancer. Therefore, the discovery of new chemical entities with the ability
to overcome the drug resistance in tumor cells is likely to improve the therapeutic index. To
investigate whether the thiazole-tethered DHAA derivatives synthesized above were still effective
on drug resistant cancer cell lines, the most potent derivatives 20, 21, and 24 were selected for
further evaluation of their potential on human breast adriamycin-resistant MCF-7 (MCF7/ADR)
cells at 0.5, 2.5, and 5 µM concentration. The results showed that all of them exhibited significant
inhibitory effect on MCF-7/ADR cells (Figure 2) and the growth inhibiton rates were even greater
than 50% at 2.5 and 5 µM. Particularly, compound 24 with a 3-(tert-butoxycarbonylamino)phenyl-
substituent on the C-4 position of the triazole ring showed the highest potency on MCF-7/ADR
cells with a submircomolar IC50 value (0.9 µM).
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Figure 2. Effect of derivatives 20, 21, 24 on MCF-7/ADR cells. MCF-7/ADR cells were treated with 20, 21 and
24 (0.5, 2.5 and 5 µM), respectively, for 72 h. The values are the mean ± S.D. of three independent experiments.
Due to the excellent antiproliferative data of 24, we evaluated its in vitro microsomal stability to
provide some guidance of drug-likeness properties. Briefly, 24 was incubated with various
microsomes
microsomes
(
RLM: rat liver microsomes, MouLM: mouse liver microsomes and DLM: dog liver
in buffer solution containing reduced nicotinamide adenine dinucleotide phosphate
)
(NADPH) regenerating system at 37 °C for an indicated time and the data were analyzed by
HPLC. As depicted in Table 3, 43.7% and 58.4% of 24 remained after 2 h incubation with
MouLM (t1/2 = 105.5 min) and RLM (t1/2 = 131.2 min) respectively. Although a relatively short
half-life (t1/2 = 69.3 min) was observed in DLM, there was still 59.0% remaining of compound 24
after 1 h incubation. Taken together, the results indicated that compound 24 exhibited a relatively
stable microsomal metabolic profile.
Table 3. In vitro microsomal metabolic stability of 24.
remaining %
Microsomes a
T1/2
30min 60min 90min 120min 150min 180min
(min)
MouLM
RLM
DLM
87.9
85.4
77.3
72.6
79.3
59.0
61.3
70.5
31.6
43.7
58.4
4.8
25.7
42.1
NDb
11.09 105.5
31.0
131.2
69.3
NDb
aRLM: rat liver microsomes, MouLM: mouse liver microsomes, DLM: dog liver microsomes,
bND: not determined.
Considering the excellent antiproliferative data and microsomal stability of 24, we conducted
some further mechanistic studies with it to determine whether its inhibition of cancer cell growth
was related to the induction of apoptosis [18].
As one of the characteristic events in late stage apoptotic cells was genomic DNA
fragmentation, which can be detected by terminal deoxynucleotiyl transferase (TDT)-mediated
dUTP nick end labeling (TUNEL), we investigated the characteristics of DNA firstly [19]. The
results shown in Figure 3 indicated that after treatment with 24 at 2.5 µM for 24 h, A549 cells
emitted light green fluorescent due to the fragmentation of genomic DNA, suggesting that
compound 24 could induce apoptosis in A549 cells. Moreover, we evaluated the morphological
characters of microtubule in A549 cells treated with 24 (Figure SI, supporting information section)
and found that compound 24 couldn’t act on microtubule.
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Figure 3. TUNNEL assay of A549 cells treated with 24 (2.5 µM) for 24 h. Cells not treated with 24 were used as
control. The intensity of DAPI and FITC was measured by spectrofluorometer.
To further confirm this property, human MDA-MB-231 cells were treated with 24 at indicated
concentrations (0.1 and 1 µM) for 24 h, and then harvested, stained with Annexin V-FITC and PI,
and analyzed by flow cytometry (Figure 4). The results indicated that 24 could induce cell
apoptosis in a dose-dependent manner. Treatment of the MDA-MB-231 cells with 0.1 and 1.0 µM
of compound 24 for 24 h resulted in 14.2% and 22.9% cell apoptosis (early and late apoptosis)
respectively. In contrast, the untreated vehicle control brought about 5.3% cell apoptosis.
Apparently, compound 24 could mediate apoptosis in cancer cells, which at least in partial
contributed to its antiproliferative effects.
Figure 4. Apoptosis ratio detection of 24 by flow cytometry. Cells were treated with compound 24 for 24 h and
cells not treated were used as control.
Previous reports have showed that dehydroabietic acid (DHAA) analogues could induce
apoptosis of cancer cells by regulating a series of apoptosis-related proteins such as caspase-3,
PARP, Bax and Bcl-2 [6h]. To elucidate the potential mechanisms contributed to apoptosis
induced by the new derivative 24, several apoptosis-related proteins were evaluated by Western
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blotting assay. As illustrated in Figure 5, low concentrations (0.25, 0.5 and 1.0 µM) of compound
24 could dose dependently down regulation the level of anti-apoptotic protein Bcl-2 and up
regulation the level of the pro-apoptotic protein Bax in human breast MDA-MB-231 cells. In
addition, the decrease of the levels of NF-κB (p65) protein expression suggested that the reduction
of Bcl-2/Bax ratio might be caused by the inhibition of NF-κB. Meanwhile, the appearance of
cleaved caspase 3 and cleaved PARP could also be viewed as markers of apoptosis. Taken
together, the present preliminary data suggested that compound 24 could induce apoptosis in
tumor cells at low concentrations. Other than apoptosis, DHAA analogues have also been reported
to inhibit tumor cell proliferation and/or induce tumor cell death through DNA binding [20], cell
cycle arrest [21], or oncosis inducing [22]. More detailed mechanism investigations on the newly
synthesized hybrid 24 are ongoing and will be disclosed in due time.
Figure 5. Western blotting analysis of biological markers for apoptosis induction by compound 24 in the MDA-
MB-231 cells (48 h). Cells not treated with 24 were used as control.
3. Conclusions
Taken together, a concise and efficient synthetic approach has been established to readily access
a series of novel C-14 1,2,3-triazole-tethered dehydroabietic acid derivatives in moderate to high
yields. In vitro antiproliferative activity evaluation of their potential against four different human
cancer cell lines, including ovary (SKOV-3), prostate (PC-3) and breast (MDA-MB-231 and
MCF-7), showed that most of the newly synthesized hybrids exhibited potent antiproliferative
activities. The SAR studies revealed that the introduction of 1,2,3-triazole moiety would be
beneficial to cytotoxicity and the substituents on the C-4 position of the triazole ring played a
critical role in exhibiting promising antiproliferative activities. Further in vitro studies
demonstrated that these new analogues, such as 20, 21, and 24 with different substituents on the
C-4 position of the triazole moiety, remained effective against adriamycin-resistant MCF-7 cells at
low concentrations in a dose-dependent manner. Particularly, derivative 24 with a 3-(tert-
butoxycarbonylamino)phenyl-substituted thiazole moiety showed the highest potency with the
IC50 values ranging from 0.7 to 1.2 µM, which are more potent than the commercial anticancer
drug 5-Fu (average IC50 value 16.1 µM). Moreover, 24 not only showed broad-spectrum
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antiproliferative activities against a series of tumor cells derived from different organs, but also
exhibited very weak cytotoxicity on normal cells, implying that there was a therapeutic window to
use compound 24. Further, preliminary mechanism studies indicated that compound 24 could
induced apoptosis in cancer cells and was worth developing into a novel natural product-like
anticancer lead by proper structure modification.
4. Experimental protocols
4.1 Biology
4.1.1 Cell lines and cell culture: Human prostate cancer PC-3, LNCaP and ovarian cancer SK-
OV-3 cell lines were from Japanese Foundation of Cancer Research (Tokyo, Japan). Human
lungadeno carcinoma A549, colorectal cancer HCT-116, breast MCF-7 and MDA-MB-231, HFF
and HL-7702 were purchased from the American Type Culture Collection (Manassas, VA).
Antibodies to cytochrome C, cleaved caspase-3, PARP, cleaved PARP, Bax, Bcl-2 and β-Actin
were purchased from Cell Signaling. All these cell lines except MCF-7 were maintained in RPMI
1640 medium (Gibco, Grand Island, NE) supplemented with 10% (v/v) heat-inactivated fetal
bovine serum (Gibco), 2 mmol/L glutamine, 100 IU/mL penicillin, 100 µg/mL streptomycin, and
10 mmol/L HEPS (pH 7.4) in a humidified atmosphere of 95% air with 5% CO2 at 37 °C. MCF-7
cells were grown in the same medium supplemented with 1 mmol/L sodium pyruvate and 0.01
mg/mL bovine insulin.
4.1.2 Whole cell lysates preparations: After treatment with compound 24, cells were washed twice
with ice-cold phosphate-buffered saline and lysed with buffer (20 mM Tris-base, pH 8.0, 100 mM
NaCl, 1 mM EDTA, 1 mM dithiothreitol, 1 mM Na3VO4, 4 µg/mL aprotinin, 1 mM phenyl-
methylsulfonyl fluoride (PMSF), 1% Triton X-100, 1% NP40, 50 mM NaF). After being
centrifugated at 12000 rpm for 15 min, the supernatant was collected as the total cellular protein
extract. Protein concentration was determined by a BCA protein assay kit.
4.2 Chemistry
4.2.1 General
All commercially available starting materials and solvents were reagent grade and were
purchased from Adamas-beta and used without further purification. Dehydroabietic acid (DHAA,
2) were purchased from Shanghai GeorGene Biotech Co. Ltd. and its purity was 98%.
Melting points were determined on a Buchi 510 melting point apparatus and were uncorrected.
Elemental analyses were performed on a Vario EL III elemental analysis instrument. Mass spectra
were obtained on Finnigan MAT Incos 50 Galaxy System (DIP-MS) (EI) or Finnigan MAT 900
(ESI) spectrometers, high resolution mass spectra on a Finnigan HSQ-30 (HR-EIMS) or on a
Finnigan MAT 900 (HR-ESI-MS). 1H and 13C NMR spectra were determined on Bruker AM-300,
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Bruker AM-400, Bruker AM-500 instruments using tetramethylsilaneas as internal reference. Data
were presented as follows: chemical shift, multiplicity (s = singlet, brs = broad singlet, d = doublet,
brd = broad doublet, t = triplet, m = multiplet), J = coupling constant in hertz (Hz). The signals of
the 13C NMR were assigned utilizing DEPT experiments and on the basis of literature data. Silica
gel 60H (200-300 mesh) manufactured by Qingdao Haiyang Chemical Group Co. (China) was
used for general chromatography.
4.2.2 Dehydroabietic acid methyl ester (3)
To a stirred solution of L-dehydroabietic acid 2 (10.0g, 33.3 mmol) and K2CO3 (5.52 g, 40 mmol)
in acetone (100 mL) was added Me2SO4 (3.47 mL, 35 mmol) dropwise and the resulting reaction
mixture was stirred at room temperature for 3 h. After removing of the solvent the mixture was
diluted with CH2Cl2, washed with brine and water, dried over anhydrous NaSO4, filtered and
concentrated under reduced pressure. Then the residue was purified by flash chromatography to
1
give 3 (10.3 g, 98.0%) as white solid, H NMR (300 MHz, CDCl3): δ 7.17 (d, J = 8.1 Hz, 1H),
6.99 (dd, J = 8.1 Hz, 1.5 Hz, 1H), 6.88 (s, 1H,), 3.67 (s, 3H), 2.91-2.83 (m, 3H), 2.32-2.22 (m, 2H),
1.83-1.64 (m, 5H), 1.54-1.48 (m, 1H), 1.39-1.36 (m, 1H,), 1.27 (s, 3H), 1.23 (s, 3H), 1.21 (s, 6H);
13C NMR (100 MHz, CDCl3):
δ 179.1, 146.9, 145.7, 134.7, 126.8, 124.1, 123.9, 51.9, 47.6, 44.8,
37.9, 36.9, 36.6, 33.4, 29.9, 25.1, 23.9, 23.8, 21.7, 18.5, 16.5; HRMS-ESI (m/z) [M + Na]+ calcd
for C21H30O2Na, 337.2143; found, 337.2149. 1H and 13C assignments matched with those
previously reported in the literature [6b].
4.2.3 12-nitrodehydroabietic acid methyl ester (4) and 14-nitrodehydroabietic acid methyl ester (5)
A solution of 68% nitric acid (1.2 ml, 19.4 mmol) in Ac2O (5 ml) was added dropwise to a stirred
solution of 3 (3.05 g, 9.7 mmol) in Ac2O (15 ml) at 0 °C over a period of 30 min, and the resulting
reaction mixture was stirred at 0 °C for another 30 min. Then the reaction mixture was poured into
an ice bath and extracted with EtOAc. The organic layer was washed successively with 1 N NaOH,
saturated NaHCO3, brine and water, dried over anhydrous NaSO4, filtered and concentrated under
reduced pressure. Then the residue was purified by flash chromatography to give 4 and 5.
Compound 4: white solid (1.36 g, 3.78 mmol, 39%), 1H NMR (400 MHz, CDCl3): δ 7.63 (s, 1H),
7.08 (s, 1H), 3.67 (s, 3H), 3.41 (sept, J = 6.9 Hz, 1H), 2.96-2.91 (m, 2H), 2.29 (brd, J = 12.3 Hz,
1H), 2.18 (dd, J = 12.6, 2.1 Hz, 1H), 1.83-1.66 (m, 5H), 1.55-1.43 (m, 2H), 1.26 (s, 3H), 1.40 (d,
J=6.8 Hz, 3H), 1.28-121 (m, 12H); 13C NMR(100 MHz, CDCl3)
δ 178.7, 148.1, 147.5, 140.8,
139.6, 127.9, 120.3, 52.0, 47.3, 44.3, 37.6, 37.0, 36.5, 29.7, 28.0, 24.9, 23.6, 23.5, 21.1, 18.2, 16.4;
HRMS-ESI (m/z) [M + Na]+ calcd for C21H29NO4Na, 382.1994; found, 382.1997.
1
Compound 5: white solid (697.4 mg, 1.94 mmol, 20%), H NMR (400 MHz, CDCl3)
δ 7.34 (d, J
= 8.6 Hz, 1H),7.18 (d, J = 8.4 Hz, 1H), 3.67 (s, 3H), 2.83-2.66 (m, 3H), 2.29 (brd, J = 11.1 Hz,
1H), 2.20 (dd, J = 12.6 Hz, 2.1 Hz, 1H), 1.83-1.63 (m, 5H), 1.52-1.46 (m, 2H), 1.26 (s, 3H), 1.5-
12

ACCEPTED MANUSCRIPT
1.19 (m, 9H); 13C NMR (100 MHz, CDCl3)
δ
178.6, 150.9, 148.9, 136.2, 126.2, 125.2, 123.9, 52.1,
47.4, 43.9, 37.9, 37.3, 36.4, 28.7, 24.9, 24.5, 23.7, 23.5, 20.5, 18.4, 16.4; HRMS-ESI (m/z) [M +
1
Na]+ calcd for C21H29NO4Na, 382.1994; found, 382.1999. H and 13C assignments matched with
those previously reported in the literature [23].
4.2.4 14-Aminodehydroabietic acid methyl ester (6)
To a stirred solution of 5 (3.6 g, 10 mmol) and NH4Cl (3.21 g, 60 mmol) in ethanol (50 mL) and
water (5 ml) was added Fe dust (2.79 g, 50 mmol). The resulting reaction mixture was heated
under reflux for 3 h. After cooled to room temperature, the solid was removed by filtration
through a pad of Celite and the solvent was removed under reduced pressure. The crude product
was purified by flash chromatography to give 6 (2.47 g, 7.5 mmol, 75%) as white solid, 1H NMR
(500 MHz, Chloroform-d) δ 7.02 (d, J = 8.3 Hz, 1H), 6.77 (d, J = 8.3 Hz, 1H), 3.67 (s, 3H), 3.62
(s, 2H), 2.89 (hept, J = 6.8 Hz, 1H), 2.66 – 2.49 (m, 2H), 2.34 – 2.29 (m, 1H), 2.26 (dd, J = 12.8,
2.1 Hz, 1H), 1.88 (tdd, J = 13.0, 11.0, 7.2 Hz, 1H), 1.81 – 1.61 (m, 4H), 1.58 – 1.45 (m, 2H), 1.29
(s, 3H), 1.29 (d, J = 7.6 Hz, 3H), 1.26 (d, J = 6.8 Hz, 3H), 1.24 (s, 3H); 13C NMR (125 MHz,
CDCl3) δ 179.2, 147.9, 140.7, 128.8, 122.7, 119.5, 114.3, 51.9, 47.6, 44.1, 38.2, 37.0, 36.6, 27.7,
25.7, 25.1, 22.5, 22.2, 21.4, 18.7, 16.5; HRMS-ESI (m/z) [M + Na]+ calcd for C21H31NO2Na,
352.2252; found, 352.2248. 1H and 13C assignments matched with those previously reported in the
literature [23].
4.2.5 14-(2-chloroacetamido)dehydroabietic acid methyl ester (7)
To a stirred solution of 6 (2.5 g, 7.59 mmol) in dry THF (120 mL) was added dropwise
chloracetyl chloride (1 M in THF, 15.2 mL, 15.2 mmol) and TEA (31.2 mL, 22.3 mmol) at 0 °C.
After stirring for 30 min at the same temperature, the mixture was warmed to room temperature
and stirred for another 2 h. The reaction mixture was diluted with water, extracted with
dichloromethane, dried over anhydrous Na2SO4, filtered, concentrated and purified by flash
1
chromatography to give 7 (2.5 g, 95 mmol, 81%) as white solid, mp 125-128 °C; H NMR (500
MHz, Chloroform-d) δ 6.78 (d, J = 8.4 Hz, 1H), 6.67 (d, J = 8.4 Hz, 1H), 3.75 (s, 2H), 3.15 (s,
3H), 2.49 (hept, J = 6.8 Hz, 1H), 2.31 – 2.06 (m, 2H), 1.81 (dd, J = 12.7, 3.0 Hz, 1H), 1.75 – 1.63
(m, 1H), 1.36 – 1.17 (m, 5H), 1.14 (d, J = 7.0 Hz, 1H), 1.07 – 0.87 (m, 2H), 0.76 (s, 3H), 0.71 (m,
6H), 0.68 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.2, 165.7, 148.2, 142.9, 132.6,
130.5, 124.9, 123.5, 52.1, 47.5, 44.0, 42.7, 38.0, 37.1, 36.4, 28.4, 26.2, 25.1, 23.7, 23.3, 21.2, 18.5,
16.5; HRMS-ESI (m/z) [M + H]+ calcd for C23H33ClNO3, 406.2149, found, 406.2160.
4.2.6 14-(2-azidoacetamido)dehydroabietic acid methyl ester (8)
To a stirred solution of 7 (2.0 g, 4.93 mmol) in DMF (20.0 mL), NaN3 ( 887 mg, 14.78 mmol) was
added, and the mixture was stirred at 80 °C for 12 h, then cooled to room temperature. After
removal of the solvent under reduced pressure, the residue was diluted with water and then
13

ACCEPTED MANUSCRIPT
extracted with ethyl acetate, washed with brine, dried with anhydrous Na2SO4 and purified by
chromatography to give 8 as white solid (1.57 g, 3.80 mmol, 77%). mp 182-184 °C; 1H NMR (500
MHz, Chloroform-d) δ 6.80 (d, J = 8.4 Hz, 1H), 6.68 (d, J = 8.4 Hz, 1H), 3.76 – 3.52 (m, 2H),
3.17 (s, 3H), 2.50 (p, J = 6.9 Hz, 1H), 2.31 – 2.08 (m, 2H), 1.88 – 1.77 (m, 1H), 1.77 – 1.69 (m,
1H), 1.28 (ddt, J = 20.0, 14.3, 7.9 Hz, 4H), 1.16 (d, J = 7.8 Hz, 1H), 1.08 – 0.91 (m, 2H), 0.80 (s,
3H), 0.74 (s, 3H), 0.72 (d, J = 6.9 Hz, 3H), 0.68 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3)
δ 179.0, 165.9, 148.0, 142.9, 132.6, 130.9, 124.6, 123.4, 52.6, 52.0, 47.5, 44.1, 38.0, 37.1, 36.4,
28.4, 26.3, 25.2, 23.7, 23.3, 21.2, 18.6, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C23H33N4O3,
413.2553, found, 413.2559.
4.2.7 General procedure for the synthesis of compounds 9-33
A solution of the azide 12 (0.1 mmol), the alkyne (0.12 mmol), 1 M aq CuSO4·5H2O (0.02 mmol)
and 1 M aq. sodium ascorbate (0.02 mmol) in a t-BuOH-H2O 1:1 mixture (3.0 mL) was heated at
50 °C until TLC showed no traces of the starting azide. The solvent was removed under reduced
pressure, the residue was dispersed in EtOAc (10 mL) and the solution was washed with H2O,
dried over MgSO4, concentrated and purified by chromatography on silica gel to give the target
analogue.
4.2.7.1 14-(2-(4,5-Bis(methoxycarbonyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid
methyl ester(9)
White solid, (39.9 mg, 72%), mp 203-205 °C; 1H NMR (500 MHz, Chloroform-d) δ 7.73 (s, 1H),
7.20 (d, J = 8.5 Hz, 1H), 7.09 (d, J = 8.4 Hz, 1H), 5.61 (s, 2H), 3.93 (s, 3H), 3.87 (s, 3H), 3.61 (s,
3H), 2.93 (h, J = 7.1 Hz, 1H), 2.73 – 2.52 (m, 2H), 2.25 (d, J = 12.7 Hz, 1H), 2.12 (d, J = 12.2 Hz,
1H), 1.70 (m, 4H), 1.62 – 1.57 (m, 1H), 1.45 – 1.35 (m, 2H), 1.23 (s, 3H), 1.15 (s, 3H), 1.10 (d, J
= 6.9 Hz, 3H), 1.08 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.1, 163.3, 160.1, 159.0,
148.0, 143.1, 139.6, 132.7, 131.2, 130.7, 124.7, 123.4, 53.5, 52.7, 52.6, 52.0, 47.5, 44.1, 44.0, 38.0,
37.1, 36.5, 28.2, 26.2, 23.7, 23.3, 21.2, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for
C29H39N4O7, 555.2819, found, 555.2871; Anal. Calc. for C29H38N4O7: C, 62.80; H, 6.91; N, 10.10;
O, 20.19. Found: C, 62.71; H, 6.99; N, 10.04; O, 20.30.
4.2.7.2 14-(2-(4-(bromomethyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(10)
White solid, (32.4 mg, 61%), mp 193-196 °C; 1H NMR (500 MHz, Chloroform-d) δ 7.80 (s, 1H),
7.49 (s, 1H), 7.22 (d, J = 8.5 Hz, 1H), 7.10 (d, J = 8.3 Hz, 1H), 5.23 (s, 2H), 4.77 (s, 2H), 3.63 (s,
3H), 2.85 (p, J = 6.8 Hz, 1H), 2.59 (d, J = 7.7 Hz, 2H), 2.26 (d, J = 12.9 Hz, 1H), 2.14 (d, J = 12.5
Hz, 1H), 1.71 (td, J = 18.5, 15.5, 7.9 Hz, 4H), 1.63 (d, J = 8.5 Hz, 1H), 1.48 – 1.35 (m, 2H), 1.24
(s, 3H), 1.17 (s, 3H), 1.12 (d, J = 6.8 Hz, 3H), 1.09 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz,
CDCl3) δ 179.2, 164.3, 148.2, 142.9, 132.5, 130.5, 129.9, 124.8, 123.4, 53.3, 52.1, 47.5, 44.1, 38.0,
14

ACCEPTED MANUSCRIPT
37.1, 36.5, 28.4, 27.2, 26.1, 25.1, 23.6, 23.2, 21.2, 18.5, 16.4; HRMS-ESI (m/z) [M + H]+ calcd for
C26H36BrN4O3, 531.1971, found, 531.1978; Anal. Calc. for C26H35BrN4O3: C, 58.76; H, 6.64; N,
10.54; O, 9.03. Found: C, 58.91; H, 6.73; N, 10.47; O, 9.09.
4.2.7.3 14-(2-(4-cyclopropyl-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl ester(11)
White solid, (21.5 mg, 45%), mp 201-204 °C; 1H NMR (500 MHz, Chloroform-d) δ 7.50 (s, 1H),
7.27 (s, 1H), 7.22 (d, J = 8.4 Hz, 1H), 7.09 (d, J = 8.4 Hz, 1H), 5.17 (s, 2H), 3.64 (s, 3H), 2.78
(hept, J = 6.8 Hz, 1H), 2.61 – 2.46 (m, 2H), 2.30 – 2.22 (m, 1H), 2.14 (d, J = 12.5 Hz, 1H), 1.99 (q,
J = 7.9, 6.7 Hz, 1H), 1.80 – 1.65 (m, 4H), 1.62 – 1.58 (m, 1H), 1.44 (dt, J = 15.0, 7.3 Hz, 1H),
1.39 – 1.33 (m, 1H), 1.23 (s, 3H), 1.16 (s, 3H), 1.11 (d, J = 6.9 Hz, 3H), 1.07 (d, J = 6.9 Hz, 3H),
0.99 (p, J = 3.9, 3.1 Hz, 2H), 0.89 – 0.83 (m, 2H); 13C NMR (125 MHz, CDCl3) δ 179.0, 164.4,
148.2, 142.7, 132.5, 130.4, 124.8, 123.4, 53.4, 52.0, 47.5, 44.0, 38.0, 37.1, 36.4, 28.4, 26.1, 25.2,
23.6, 23.2, 21.1, 18.5, 16.5, 8.0, 8.0, 6.6; HRMS-ESI (m/z) [M + H]+ calcd for C28H39N4O3,
479.3022, found, 479.3032; Anal. Calc. for C28H38N4O3: C, 70.26; H, 8.00; N, 11.71; O, 10.03.
Found: C, 70.11; H, 8.15; N, 11.64; O, 10.13.
4.2.7.4 14-(2-(4-(ethoxycarbonyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(12)
White solid, (32.2 mg, 63%), mp 189-192 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.41 (s, 1H),
8.15 (s, 1H), 7.20 (d, J = 8.4 Hz, 1H), 7.08 (d, J = 8.3 Hz, 1H), 5.37 (d, J = 4.9 Hz, 2H), 4.35 (q, J
= 7.1 Hz, 2H), 3.70 – 3.64 (m, 1H), 3.61 (s, 3H), 2.87 (h, J = 6.8 Hz, 1H), 2.66 – 2.54 (m, 2H),
2.25 (d, J = 12.9 Hz, 1H), 2.11 (d, J = 12.5 Hz, 1H), 1.79 – 1.56 (m, 5H), 1.33 (t, J = 7.1 Hz, 4H),
1.22 (s, 3H), 1.14 (s, 3H), 1.07 (d, J = 6.7 Hz, 3H), 1.04 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz,
CDCl3) δ 179.22, 163.70, 160.49, 148.04, 142.92, 140.41, 132.59, 130.73, 129.35, 124.69, 123.36,
61.46, 58.35, 52.88, 52.05, 47.51, 44.08, 37.05, 28.32, 26.11, 25.11, 23.59, 23.19, 21.17, 18.50,
18.38, 16.44, 14.27; HRMS-ESI (m/z) [M + H]+ calcd for C28H39N4O5, 511.2920, found, 511.2924;
Anal. Calc. for C28H38N4O5: C, 65.86; H, 7.50; N, 10.97; O, 15.67. Found: C, 65.95; H, 7.63; N,
10.99; O, 15.79.
4.2.7.5
14-(2-(4-((trimethylsilyloxy)carbonyl)-1H-1,2,3-triazol-1-yl)acetamidodehydro)abietic
acid methyl ester(13)
White solid, (23.5 mg, 46%), mp 155-157 °C; 1H NMR (500 MHz, Chloroform-d) δ 7.82 (s, 1H),
7.25 (d, J = 8.4 Hz, 1H), 7.12 (d, J = 8.4 Hz, 1H), 5.34 (s, 2H), 3.66 (s, 3H), 2.79 (hept, J = 6.8 Hz,
1H), 2.63 – 2.48 (m, 2H), 2.32 – 2.26 (m, 1H), 2.18 (d, J = 12.6 Hz, 1H), 1.74 (ddd, J = 18.5, 9.0,
5.2 Hz, 4H), 1.63 (d, J = 7.7 Hz, 1H), 1.48 (d, J = 11.4 Hz, 1H), 1.38 (dd, J = 12.8, 6.7 Hz, 1H),
1.26 (s, 3H), 1.19 (s, 3H), 1.13 (d, J = 6.8 Hz, 3H), 1.10 (d, J = 6.9 Hz, 3H), 0.38 (s, 9H); 13C
NMR (125 MHz, CDCl3) δ 178.9, 164.4, 148.2, 142.7, 132.5, 130.4, 129.9, 124.8, 123.4, 53.0,
52.0, 47.5, 44.0, 38.0, 37.1, 36.4, 28.4, 26.1, 25.1, 23.5, 23.1, 21.1, 18.5, 16.5, -1.2; HRMS-ESI
15

ACCEPTED MANUSCRIPT
(m/z) [M + H]+ calcd for C28H43N4O3Si, 511.3104, found, 511.3123; Anal. Calc. for C28H42N4O3Si:
C, 65.85; H, 8.29; N, 10.97; O, 9.40. Found: C, 65.88; H, 8.35; N, 10.78; O, 9.61.
4.2.7.6 14-(2-(4-benzyl-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methylester(14)
White solid, (41.8 mg, 79%), mp 188-190 °C; 1H NMR (500 MHz, Chloroform-d) δ 7.44 (s, 1H),
7.36 – 7.30 (m, 2H), 7.29 – 7.22 (m, 4H), 7.11 (d, J = 8.3 Hz, 1H), 5.18 (s, 2H), 4.14 (s, 2H), 3.66
(s, 3H), 2.79 (hept, J = 6.9 Hz, 1H), 2.55 (dd, J = 9.2, 4.6 Hz, 2H), 2.29 (d, J = 12.1 Hz, 1H), 2.17
(d, J = 12.5 Hz, 1H), 1.75 (tdd, J = 13.7, 7.3, 4.0 Hz, 4H), 1.64 (d, J = 6.8 Hz, 1H), 1.46 (dd, J =
13.8, 9.9 Hz, 1H), 1.37 (dt, J = 11.9, 3.9 Hz, 1H), 1.27 (s, 3H), 1.18 (s, 3H), 1.11 (d, J = 6.9 Hz,
3H), 1.07 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.0, 164.3, 148.2, 142.8, 138.5,
132.5, 130.4, 128.8, 128.7, 126.8, 124.8, 123.4, 53.3, 52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 32.2, 28.4,
26.1, 25.1, 23.5, 23.1, 21.1, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C32H41N4O3,
529.3179, found, 529.3187; Anal. Calc. for C32H40N4O3: C, 72.70; H, 7.63; N, 10.60; O, 9.08.
Found: C, 72.90; H, 7.78; N, 10.69; O, 9.15.
4.2.7.7 14-(2-(4-phenyl-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl ester(15)
White solid, (43.8 mg, 85%), mp 211-214 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.06 (s, 1H),
7.84 – 7.79 (m, 2H), 7.57 (s, 1H), 7.43 (dd, J = 8.3, 6.8 Hz, 2H), 7.39 – 7.33 (m, 1H), 7.27 – 7.19
(m, 1H), 7.10 (d, J = 8.4 Hz, 1H), 5.30 (s, 2H), 3.60 (s, 3H), 2.85 (p, J = 6.8 Hz, 1H), 2.67 – 2.52
(m, 2H), 2.26 (dd, J = 12.0, 3.3 Hz, 1H), 2.14 (d, J = 12.5 Hz, 1H), 1.83 (s, 1H), 1.77 – 1.63 (m,
4H), 1.60 (q, J = 3.0 Hz, 1H), 1.39 – 1.32 (m, 1H), 1.22 (s, 3H), 1.15 (s, 3H), 1.10 (d, J = 7.0 Hz,
3H), 1.08 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.0, 164.3, 148.2, 142.8, 132.6,
130.4, 129.8, 129.0, 128.7, 125.8, 124.9, 123.4, 121.3, 53.4, 52.0, 47.5, 44.0, 38.0, 37.1, 36.4, 28.4,
26.2, 25.2, 23.6, 23.2, 21.2, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C31H39N4O3,
515.3022, found, 515.3031; Anal. Calc. for C31H38N4O3: C, 72.35; H, 7.44; N, 10.89; O, 9.33.
Found: C, 72.42; H, 7.51; N, 10.80; O, 9.44.
4.2.7.8 14-(2-(4-(2-fluorophenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(16)
White solid, (39.9 mg, 75%), mp 153-156 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.27 (td, J =
7.6, 1.7 Hz, 1H), 8.22 (d, J = 3.4 Hz, 1H), 7.42 (s, 1H), 7.39 – 7.33 (m, 1H), 7.28 (t, J = 3.8 Hz,
1H), 7.24 (d, J = 8.3 Hz, 1H), 7.18 (dd, J = 11.0, 8.4 Hz, 1H), 7.12 (d, J = 8.3 Hz, 1H), 5.34 (s,
2H), 3.64 (s, 3H), 2.88 (p, J = 6.9 Hz, 1H), 2.69 – 2.53 (m, 2H), 2.32 – 2.22 (m, 1H), 2.17 (d, J =
12.5 Hz, 1H), 1.80 – 1.67 (m, 4H), 1.63 (d, J = 6.9 Hz, 1H), 1.47 (dd, J = 12.6, 3.6 Hz, 1H), 1.41 –
1.35 (m, 1H), 1.25 (s, 3H), 1.18 (s, 3H), 1.13 (d, J = 7.1 Hz, 3H), 1.11 (d, J = 7.0 Hz, 3H); 13C
NMR (125 MHz, CDCl3) δ 179.0, 164.2, 160.3, 158.4, 148.2, 142.8, 142.2, 132.6, 130.4, 129.9,
129.9, 127.8, 127.8, 124.9, 124.7, 124.7, 124.3, 124.2, 123.4, 117.9, 117.8, 115.9, 115.7, 53.4,
52.0, 47.5, 44.0, 38.0, 37.1, 36.4, 28.4, 26.2, 25.1, 23.5, 23.2, 21.1, 18.5, 16.5; HRMS-ESI (m/z)
16

ACCEPTED MANUSCRIPT
[M + H]+ calcd for C31H38FN4O3, 533.2928, found, 533.2934; Anal. Calc. for C31H37FN4O3: C,
69.90; H, 7.00; N, 10.52; O, 9.01. Found: C, 69.99; H, 7.17; N, 10.40; O, 9.15.
4.2.7.9 14-(2-(4-(3-fluorophenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(17)
White solid, (42.6 mg, 80%), mp 164-167 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.10 (s, 1H),
7.60 (q, J = 11.6, 8.3 Hz, 2H), 7.52 (d, J = 4.4 Hz, 1H), 7.41 (dt, J = 15.7, 8.1 Hz, 1H), 7.26 (dd, J
= 8.7, 4.8 Hz, 1H), 7.13 (dd, J = 8.5, 4.4 Hz, 1H), 7.08 (td, J = 8.5, 3.2 Hz, 1H), 5.33 (d, J = 4.8
Hz, 2H), 3.64 (s, 3H), 2.92 – 2.82 (m, 1H), 2.62 (q, J = 11.3, 7.6 Hz, 2H), 2.29 (d, J = 12.8 Hz,
1H), 2.16 (d, J = 12.5 Hz, 1H), 1.80-1.66 (t, J = 12.1 Hz, 4H), 1.66 – 1.59 (m, 1H), 1.50 – 1.35 (m,
2H), 1.25 (s, 3H), 1.18 (s, 3H), 1.13 (d, J = 6.6 Hz, 3H), 1.10 (s, 3H); 13C NMR (125 MHz,
CDCl3) δ 179.0, 164.1, 162.2, 148.2, 142.8, 132.5, 130.7, 130.6, 130.4, 124.9, 123.5, 121.5, 115.6,
115.4, 112.9, 112.7, 53.4, 52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 28.4, 26.2, 25.1, 23.6, 23.2, 21.2, 18.5,
16.5; HRMS-ESI (m/z) [M + H]+ calcd for C31H38FN4O3, 533.2928, found, 533.2941; Anal. Calc.
for C31H37FN4O3: C, 69.90; H, 7.00; N, 10.52; O, 9.01. Found: C, 70.10; H, 7.13; N, 10.41; O,
9.10.
4.2.7.10 14-(2-(4-(4-fluorophenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(18)
White solid, (44.2 mg, 83%), mp 172-175 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.03 (s, 1H),
7.81 – 7.74 (m, 2H), 7.61 (s, 1H), 7.23 (d, J = 8.4 Hz, 1H), 7.11 (m, 3H), 5.30 (s, 2H), 3.61 (s, 3H),
2.85 (hept, J = 6.7 Hz, 1H), 2.61 (qd, J = 13.6, 11.5, 7.1 Hz, 2H), 2.26 (d, J = 12.7 Hz, 1H), 2.13
(d, J = 12.5 Hz, 1H), 1.70 (td, J = 11.3, 4.7 Hz, 4H), 1.64 – 1.58 (m, 1H), 1.43 (d, J = 12.2 Hz,
1H), 1.36 (dd, J = 12.7, 6.1 Hz, 1H), 1.22 (s, 3H), 1.15 (s, 3H), 1.10 (d, J = 6.8 Hz, 3H), 1.07 (d, J
= 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.1, 164.2, 163.9, 161.9, 148.2, 147.8, 142.8,
132.5, 130.4, 127.6, 126.1, 124.9, 123.5, 121.1, 116.1, 115.9, 53.4, 52.0, 47.5, 44.0, 38.0, 37.1,
36.5, 28.4, 26.2, 25.2, 23.6, 23.2, 21.2, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for
C31H38FN4O3, 533.2928, found, 533.2939; Anal. Calc. for C31H37FN4O3: C, 69.90; H, 7.00; N,
10.52; O, 9.01. Found: C, 70.02; H, 7.10; N, 10.47; O, 9.14.
5.2.7.11 14-(2-(4-(2-methoxyphenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(19)
White solid, (39.8 mg, 73%), mp 193-195 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.33 (dd, J =
7.7, 1.7 Hz, 1H), 8.29 (s, 1H), 7.36 (ddd, J = 8.8, 7.5, 1.7 Hz, 1H), 7.25 (s, 1H), 7.22 (d, J = 8.4
Hz, 1H), 7.13 – 7.05 (m, 2H), 7.01 (d, J = 8.3 Hz, 1H), 5.30 (s, 2H), 3.96 (s, 3H), 3.60 (s, 3H),
2.84 (hept, J = 6.9 Hz, 1H), 2.60 (q, J = 9.4, 8.6 Hz, 2H), 2.29 – 2.18 (m, 1H), 2.15 (d, J = 12.4 Hz,
1H), 1.77 – 1.66 (m, 4H), 1.60 (d, J = 7.2 Hz, 1H), 1.45 (d, J = 12.0 Hz, 1H), 1.37 – 1.31 (m, 1H),
1.22 (s, 3H), 1.16 (s, 3H), 1.11 (d, J = 7.8 Hz, 3H), 1.09 (d, J = 7.1 Hz, 3H); 13C NMR (125 MHz,
17

ACCEPTED MANUSCRIPT
CDCl3) δ 179.0, 164.6, 155.8, 148.1, 144.2, 142.8, 132.6, 130.4, 129.5, 127.7, 124.9, 124.6, 123.4,
121.1, 118.5, 110.8, 55.5, 53.4, 52.0, 47.5, 43.9, 38.0, 37.1, 36.4, 28.4, 26.2, 25.2, 23.6, 23.2, 21.1,
18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C32H41N4O4, 545.3128, found, 545.3135; Anal.
Calc. for C32H40N4O4: C, 70.56; H, 7.40; N, 10.29; O, 11.75. Found: C, 70.39; H, 7.45; N, 10.33;
O, 11.84.
4.2.7.12 14-(2-(4-(3-methoxyphenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(20)
White solid, (45.8 mg, 84%), mp 175-177 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.06 (s, 1H),
7.67 (d, J = 6.1 Hz, 1H), 7.44 – 7.40 (m, 1H), 7.36 – 7.29 (m, 2H), 7.22 (d, J = 8.4 Hz, 1H), 7.09
(d, J = 8.4 Hz, 1H), 6.90 (dt, J = 7.6, 2.1 Hz, 1H), 5.29 (s, 2H), 3.84 (s, 3H), 3.60 (s, 3H), 2.86
(hept, J = 7.1 Hz, 1H), 2.65 – 2.53 (m, 2H), 2.25 (d, J = 12.3 Hz, 1H), 2.13 (d, J = 12.5 Hz, 1H),
1.77 – 1.63 (m, 4H), 1.63 – 1.57 (m, 1H), 1.43 (d, J = 11.7 Hz, 1H), 1.35 (dt, J = 12.5, 6.7 Hz, 1H),
1.22 (s, 3H), 1.15 (s, 3H), 1.10 (d, J = 6.9 Hz, 3H), 1.07 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz,
CDCl3) δ 179.1, 164.3, 160.1, 148.5, 148.1, 142.8, 132.5, 131.2, 130.5, 130.0, 124.8, 123.4, 121.6,
118.2, 114.6, 110.9, 55.4, 53.3, 52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 28.4, 26.2, 25.1, 23.6, 23.2, 21.2,
18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C32H41N4O4, 545.3128, found, 545.3129; Anal.
Calc. for C32H40N4O4: C, 70.56; H, 7.40; N, 10.29; O, 11.75. Found: C, 70.70; H, 7.62; N, 10.39;
O, 11.90.
4.2.7.13 14-(2-(4-(4-methoxyphenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(21)
White solid, (47.9 mg, 88%), mp 194-196 °C; 1H NMR (500 MHz, Chloroform-d) δ 7.96 (s, 1H),
7.73 (dd, J = 8.8, 2.4 Hz, 2H), 7.61 (d, J = 13.5 Hz, 1H), 7.22 (d, J = 8.4 Hz, 1H), 7.10 (d, J = 8.4
Hz, 1H), 6.98 – 6.92 (m, 2H), 5.29 (s, 2H), 3.84 (s, 3H), 3.61 (s, 3H), 2.85 (p, J = 6.8 Hz, 1H),
2.68 – 2.50 (m, 2H), 2.26 (d, J = 12.7 Hz, 1H), 2.14 (d, J = 12.4 Hz, 1H), 1.70 (td, J = 11.7, 6.2
Hz, 4H), 1.60 (d, J = 6.6 Hz, 1H), 1.43 (t, J = 12.0 Hz, 1H), 1.35 (dd, J = 13.0, 6.6 Hz, 1H), 1.22
(s, 3H), 1.15 (s, 3H), 1.10 (d, J = 6.9 Hz, 3H), 1.07 (d, J = 6.8 Hz, 3H); 13C NMR (125 MHz,
CDCl3) δ 179.0, 164.3, 159.9, 148.5, 148.1, 142.8, 132.5, 130.5, 127.2, 124.8, 123.4, 122.5, 120.5,
114.4, 55.3, 53.5, 53.3, 52.0, 47.5, 44.0, 38.0, 37.1, 36.4, 28.4, 26.1, 25.1, 23.5, 23.2, 21.1, 18.5,
16.4; HRMS-ESI (m/z) [M + H]+ calcd for C32H41N4O4, 545.3128, found, 545.3129; Anal. Calc.
for C32H40N4O4: C, 70.56; H, 7.40; N, 10.29; O, 11.75. Found: C, 70.65; H, 7.57; N, 10.33; O,
11.92.
4.2.7.14
14-(2-(4-(4-(trifluoromethoxy)phenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic
acid methyl ester(22)
White solid, (41.3 mg, 69%), mp 169-172 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.07 (s, 1H),
7.84 (d, J = 8.3 Hz, 2H), 7.51 (d, J = 3.8 Hz, 1H), 7.28 (d, J = 8.3 Hz, 2H), 7.23 (d, J = 8.3 Hz,
18

ACCEPTED MANUSCRIPT
1H), 7.11 (d, J = 8.3 Hz, 1H), 5.31 (s, 2H), 3.61 (s, 3H), 2.85 (h, J = 7.1 Hz, 1H), 2.66 – 2.53 (m,
2H), 2.26 (d, J = 12.7 Hz, 1H), 2.14 (d, J = 12.5 Hz, 1H), 1.80 – 1.66 (m, 4H), 1.64 – 1.58 (m, 1H),
1.46 – 1.33 (m, 2H), 1.23 (s, 3H), 1.16 (s, 3H), 1.11 (d, J = 6.9 Hz, 3H), 1.08 (d, J = 6.9 Hz, 3H);
13C NMR (125 MHz, CDCl3) δ 179.0, 164.1, 149.3, 148.2, 142.8, 132.5, 130.4, 128.6, 127.3,
127.2, 124.9, 123.5, 121.5, 121.5, 119.4, 53.5, 53.4, 52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 28.5, 26.2,
25.1, 23.6, 23.2, 21.2, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C32H38F3N4O4, 599.2845,
found, 599.2881; Anal. Calc. for C32H38F3N4O4: C, 64.20; H, 6.23; N, 9.36; O, 10.69. Found: C,
64.33; H, 6.35; N, 9.31; O, 10.84.
4.2.7.15
14-(2-(4-(3-acetamidophenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic
acid
methyl ester(23)
1
White solid, (28.0 mg, 49%), mp 203-205 °C; H NMR (500 MHz, DMSO-d6) δ 10.07 (s, 1H),
9.61 (s, 1H), 8.52 (s, 1H), 8.16 (d, J = 1.9 Hz, 1H), 7.55 (dd, J = 8.0, 2.0 Hz, 1H), 7.49 (d, J = 7.7
Hz, 1H), 7.36 (t, J = 7.9 Hz, 1H), 7.24 (d, J = 8.3 Hz, 1H), 7.12 (d, J = 8.3 Hz, 1H), 5.40 (s, 2H),
3.60 (s, 3H), 3.02 (s, 1H), 2.82 (s, 1H), 2.65 (d, J = 11.7 Hz, 1H), 2.32 (d, J = 12.8 Hz, 1H), 2.06
(s, 3H), 2.01 – 1.92 (m, 1H), 1.80 – 1.53 (m, 5H), 1.36 – 1.27 (m, 2H), 1.20 (s, 3H), 1.14 (s, 3H),
1.11 (s, 3H), 1.07 (s, 3H); 13C NMR (125 MHz, DMSO-d6) δ 178.5, 168.9, 165.2, 147.9, 147.6,
146.5, 143.2, 140.3, 137.5, 131.6, 129.8, 124.2, 123.5, 123.4, 120.5, 118.9, 116.0, 52.4, 52.2, 47.4,
44.8, 43.8, 37.1, 36.6, 29.3, 28.0, 25.9, 25.2, 24.5, 21.2, 20.3, 18.5, 16.7; HRMS-ESI (m/z) [M +
H]+ calcd for C33H42N5O4, 572.3237, found, 572.3239; Anal. Calc. for C33H41N5O4: C, 69.33; H,
7.23; N, 12.25; O, 11.19. Found: C, 69.52; H, 7.32; N, 12.13; O, 11.25.
4.2.7.16
14-(2-(4-(3-(tert-butoxycarbonylamino)phenyl)-1H-1,2,3-triazol-1-
yl)acetamido)dehydroabietic acid methyl ester(24)
White solid, (44.7 mg, 71%), mp 163-166 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.06 (s, 1H),
7.87 (t, J = 1.8 Hz, 1H), 7.67 (s, 1H), 7.47 (d, J = 7.4 Hz, 1H), 7.40 – 7.28 (m, 2H), 7.22 (d, J =
8.4 Hz, 1H), 7.10 (d, J = 8.4 Hz, 1H), 6.76 (s, 1H), 5.30 (s, 2H), 3.61 (s, 3H), 3.86 (m, 1H), 2.66 –
2.53 (m, 2H), 2.26 (d, J = 12.2 Hz, 1H), 2.13 (d, J = 12.4 Hz, 1H), 1.76 – 1.63 (m, 4H), 1.60 (d, J
= 6.7 Hz, 1H), 1.52 (s, 9H), 1.38 (ddd, J = 28.0, 13.7, 5.4 Hz, 2H), 1.22 (s, 3H), 1.15 (s, 3H), 1.10
(d, J = 7.0 Hz, 3H), 1.07 (d, J = 7.0 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.0, 164.3, 152.8,
148.3, 148.1, 142.9, 139.1, 132.6, 130.6, 130.6, 129.6, 124.8, 123.4, 121.6, 120.4, 118.6, 53.3,
52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 28.4, 28.4, 26.1, 25.1, 23.6, 23.2, 21.1, 18.5, 16.5; HRMS-ESI
(m/z) [M + H]+ calcd for C36H48N5O5, 630.3655, found, 630.3663; Anal. Calc. for C36H47N5O5: C,
68.66; H, 7.52; N, 11.12; O, 12.70. Found: C, 68.75; H, 7.61; N, 11.10; O, 12.83.
4.2.7.17 14-(2-(4-(3-nitrophenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(25)
19

ACCEPTED MANUSCRIPT
1
White solid, (41.9 mg, 75%), mp 201-203 °C; H NMR (500 MHz, Chloroform-d) δ 8.64 (t, J =
2.0 Hz, 1H), 8.24 (s, 1H), 8.19 (dd, J = 8.1, 1.9 Hz, 2H), 7.62 (t, J = 8.0 Hz, 1H), 7.52 (s, 1H),
7.23 (d, J = 8.3 Hz, 1H), 7.11 (d, J = 8.2 Hz, 1H), 5.34 (s, 2H), 3.61 (s, 3H), 2.88 (p, J = 6.8 Hz,
1H), 2.62 (dd, J = 9.9, 6.8 Hz, 2H), 2.26 (d, J = 12.9 Hz, 1H), 2.13 (d, J = 12.1 Hz, 1H), 1.77 –
1.66 (m, 2H), 1.61 (dd, J = 9.3, 3.5 Hz, 1H), 1.45 – 1.35 (m, 2H), 1.22 (s, 3H), 1.15 (s, 3H), 1.12
(d, J = 6.6 Hz, 3H), 1.09 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.1, 163.9, 148.7,
148.2, 146.3, 142.8, 132.5, 131.7, 131.5, 130.4, 130.1, 125.0, 123.5, 123.2, 122.3, 120.6, 53.3,
52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 28.5, 26.2, 25.1, 23.6, 23.2, 21.2, 18.5, 16.5; HRMS-ESI (m/z)
[M + H]+ calcd for C31H38N5O5, 560.2873, found, 560.2875; Anal. Calc. for C31H37N5O5: C, 66.53;
H, 6.66; N, 12.51; O, 14.29. Found: C, 66.66; H, 6.79; N, 12.54; O, 14.45.
4.2.7.18 14-(2-(4-(4-nitrophenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(26)
1
White solid, (47.0 mg, 84%), mp 194-196 °C; H NMR (500 MHz, Chloroform-d) δ 8.34 – 8.24
(m, 3H), 8.00 (d, J = 8.6 Hz, 2H), 7.65 (s, 1H), 7.26 (d, J = 8.4 Hz, 1H), 7.14 (d, J = 8.4 Hz, 1H),
5.37 (s, 2H), 3.63 (s, 3H), 2.90 (p, J = 6.8 Hz, 1H), 2.71 – 2.58 (m, 2H), 2.29 (d, J = 12.7 Hz, 1H),
2.15 (d, J = 12.4 Hz, 1H), 1.73 (dt, J = 20.9, 6.8 Hz, 4H), 1.64 (dd, J = 10.0, 3.2 Hz, 1H), 1.47 –
1.37 (m, 2H), 1.25 (s, 3H), 1.18 (s, 3H), 1.14 (d, J = 6.9 Hz, 3H), 1.10 (d, J = 6.9 Hz, 3H); 13C
NMR (125 MHz, CDCl3) δ 179.1, 163.9, 148.3, 147.5, 146.3, 142.8, 136.2, 132.5, 130.4, 126.3,
126.2, 125.0, 124.4, 124.3, 123.5, 123.0, 53.2, 52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 28.5, 26.2, 25.2,
23.6, 23.2, 21.2, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C31H38N5O5, 560.2873, found,
560.2877; Anal. Calc. for C31H37N5O5: C, 66.53; H, 6.66; N, 12.51; O, 14.29. Found: C, 66.72; H,
6.79; N, 12.64; O, 14.38.
4.2.7.19 14-(2-(4-(3,5-dimethoxyphenyl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid
methyl ester(27)
White solid, (37.4 mg, 65%), mp 177-179 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.00 (s, 1H),
7.31 (s, 1H), 7.23 (d, J = 8.4 Hz, 1H), 7.10 (d, J = 8.4 Hz, 1H), 7.01 (d, J = 2.3 Hz, 2H), 6.48 (t, J
= 2.2 Hz, 1H), 5.29 (s, 2H), 3.85 (s, 6H), 3.62 (s, 3H), 2.83 (hept, J = 6.9 Hz, 1H), 2.58 (dd, J =
11.7, 7.1 Hz, 2H), 2.26 (d, J = 12.1 Hz, 1H), 2.15 (d, J = 12.5 Hz, 1H), 1.71 (dd, J = 18.3, 8.9 Hz,
4H), 1.61 (d, J = 7.0 Hz, 1H), 1.45 (d, J = 12.2 Hz, 1H), 1.35 (dd, J = 13.6, 8.6 Hz, 1H), 1.23 (s,
3H), 1.16 (s, 3H), 1.12 (d, J = 6.9 Hz, 3H), 1.09 (d, J = 7.0 Hz, 3H); 13C NMR (125 MHz, CDCl3)
δ 178.98, 164.11, 161.26, 148.66, 148.21, 142.75, 132.52, 131.56, 130.28, 124.92, 123.45, 121.49,
103.79, 100.95, 55.53, 53.48, 52.02, 47.51, 43.96, 37.97, 37.09, 36.44, 28.46, 26.16, 25.16, 23.58,
23.18, 21.15, 18.54, 16.47; HRMS-ESI (m/z) [M + H]+ calcd for C33H43N4O5, 575.3233, found,
575.3240; Anal. Calc. for C33H42N4O5: C, 68.97; H, 7.37; N, 9.75; O, 13.92. Found: C, 69.10; H,
7.49; N, 9.77; O, 13.99.
20

ACCEPTED MANUSCRIPT
4.2.7.20 14-(2-(4-(naphthalen-2-yl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(28)
White solid, (33.9 mg, 60%), mp 184-186 °C; 1H NMR (500 MHz, Chloroform-d) δ 8.36 (s, 1H),
8.15 (s, 1H), 7.92 (s, 1H), 7.90 – 7.84 (m, 2H), 7.51 (td, J = 7.2, 6.3, 3.4 Hz, 2H), 7.42 (s, 1H),
7.23 (d, J = 8.4 Hz, 1H), 7.11 (d, J = 8.3 Hz, 1H), 5.34 (s, 2H), 3.59 (s, 3H), 2.87 (h, J = 6.7 Hz,
1H), 2.62 (dd, J = 12.2, 7.1 Hz, 2H), 2.26 (d, J = 12.7 Hz, 1H), 2.15 (d, J = 12.6 Hz, 1H), 1.71 (dd,
J = 16.0, 7.9 Hz, 4H), 1.61 (d, J = 7.2 Hz, 1H), 1.48 – 1.34 (m, 2H), 1.22 (s, 3H), 1.16 (s, 3H),
1.12 (d, J = 6.9 Hz, 3H), 1.10 (d, J = 8.1 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.0, 164.2,
148.8, 148.2, 142.8, 133.5, 133.4, 132.6, 130.4, 128.8, 128.3, 127.8, 127.1, 126.7, 126.5, 124.9,
124.8, 123.7, 123.5, 121.5, 53.5, 52.0, 47.5, 44.0, 38.0, 37.1, 36.4, 28.5, 26.2, 25.2, 23.6, 23.2,
21.2, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C35H41N4O3, 565.3179, found, 565.3194;
Anal. Calc. for C35H40N4O3: C, 74.44; H, 7.14; N, 9.92; O, 8.50. Found: C, 74.60; H, 7.27; N,
9.99; O, 8.65.
4.2.7.21 14-(2-(4-(biphenyl-4-yl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(29)
White solid, (43.7 mg, 74%), mp 181-193 °C;1H NMR (500 MHz, Chloroform-d) δ 8.10 (s, 1H),
7.88 (d, J = 8.1 Hz, 2H), 7.65 (dt, J = 16.5, 8.3 Hz, 5H), 7.46 (t, J = 7.5 Hz, 2H), 7.40 – 7.35 (m,
1H), 7.23 (d, J = 8.4 Hz, 1H), 7.11 (d, J = 8.4 Hz, 1H), 5.33 (s, 2H), 3.61 (s, 3H), 2.88 (hept, J =
7.1 Hz, 1H), 2.70 – 2.56 (m, 2H), 2.26 (d, J = 12.3 Hz, 1H), 2.15 (d, J = 12.5 Hz, 1H), 1.72 (p, J =
12.1, 11.6 Hz, 4H), 1.64 – 1.58 (m, 1H), 1.47 – 1.35 (m, 2H), 1.23 (s, 3H), 1.16 (s, 3H), 1.12 (d, J
= 6.9 Hz, 3H), 1.09 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.0, 164.3, 148.3, 148.2,
142.9, 141.4, 140.4, 132.6, 130.5, 128.9, 128.8, 127.8, 127.7, 127.6, 127.1, 127.0, 126.2, 124.9,
123.5, 121.4, 53.5, 53.4, 52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 28.5, 26.2, 25.2, 23.6, 23.2, 21.2, 18.5,
16.5; HRMS-ESI (m/z) [M + H]+ calcd for C37H43N4O3, 591.3335, found, 591.3339; Anal. Calc.
for C37H42N4O3: C, 75.23; H, 7.17; N, 9.48; O, 8.12. Found: C, 75.40; H, 7.28; N, 9.59; O, 8.24.
4.2.7.22 14-(2-(4-(pyridin-2-yl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(30)
1
White solid, (32.5 mg, 63%), mp 174-177 °C; H NMR (500 MHz, Chloroform-d) δ 8.63 – 8.54
(m, 1H), 8.39 (s, 1H), 8.15 (d, J = 7.9 Hz, 1H), 7.80 (td, J = 7.8, 1.8 Hz, 1H), 7.49 (s, 1H), 7.28 (q,
J = 3.1 Hz, 1H), 7.23 (d, J = 8.4 Hz, 1H), 7.11 (d, J = 8.4 Hz, 1H), 5.34 (s, 2H), 3.63 (s, 3H), 2.88
(hept, J = 6.9 Hz, 1H), 2.70 – 2.51 (m, 2H), 2.34 – 2.25 (m, 1H), 2.15 (d, J = 12.7 Hz, 1H), 1.77 –
1.66 (m, 4H), 1.61 (d, J = 6.8 Hz, 1H), 1.45 (q, J = 11.3, 7.7 Hz, 1H), 1.40 – 1.34 (m, 1H), 1.23 (s,
3H), 1.16 (s, 3H), 1.12 (d, J = 7.0 Hz, 3H), 1.10 (d, J = 7.0 Hz, 3H); 13C NMR (125 MHz, CDCl3)
δ 179.0, 164.0, 149.6, 149.5, 149.2, 148.1, 142.8, 137.0, 132.6, 130.4, 124.8, 123.8, 123.4, 123.3,
120.3, 53.4, 52.0, 47.5, 44.0, 38.0, 37.1, 36.4, 28.4, 26.1, 25.2, 23.6, 23.2, 21.1, 18.5, 16.5;
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HRMS-ESI (m/z) [M + H]+ calcd for C30H38N5O3, 516.2975, found, 516.2983; Anal. Calc. for
C30H37N5O3: C, 69.88; H, 7.23; N, 13.58; O, 9.31. Found: C, 69.97; H, 7.35; N, 13.62; O, 9.38.
4.2.7.23 14-(2-(4-(pyridin-3-yl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(31)
White solid, (36.1 mg, 70%), mp 197-199 °C; 1H NMR (500 MHz, Chloroform-d) δ 9.01 (s, 1H),
8.57 (s, 1H), 8.20 (s, 1H), 8.13 (d, J = 8.0 Hz, 1H), 7.99 (d, J = 3.8 Hz, 1H), 7.35 (d, J = 9.0 Hz,
1H), 7.21 (d, J = 8.4 Hz, 1H), 7.09 (d, J = 8.3 Hz, 1H), 5.33 (s, 2H), 3.59 (s, 3H), 2.87 (hept, J =
6.7 Hz, 1H), 2.69 – 2.54 (m, 2H), 2.24 (d, J = 12.9 Hz, 1H), 2.11 (d, J = 12.4 Hz, 1H), 1.69 (td, J
= 17.3, 14.6, 7.5 Hz, 4H), 1.60 (dd, J = 9.5, 3.4 Hz, 1H), 1.37 (ddt, J = 18.4, 11.5, 7.1 Hz, 2H),
1.21 (s, 3H), 1.13 (s, 3H), 1.09 (d, J = 6.7 Hz, 3H), 1.05 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz,
CDCl3) δ 179.1, 167.2, 164.1, 149.4, 148.2, 146.9, 145.3, 142.8, 133.1, 132.5, 130.6, 124.8, 123.4,
121.9, 53.2, 52.0, 47.5, 44.1, 38.0, 37.1, 36.5, 28.4, 26.2, 25.2, 25.1, 23.6, 23.2, 21.2, 18.5, 16.4;
HRMS-ESI (m/z) [M + H]+ calcd for C30H38N5O3, 516.2975, found, 516.2987; Anal. Calc. for
C30H37N5O3: C, 69.88; H, 7.23; N, 13.58; O, 9.31. Found: C, 69.95; H, 7.33; N, 13.65; O, 9.37.
4.2.7.24 14-(2-(4-(Pyridin-4-yl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(32)
1
White solid, (27.8 mg, 54%), mp 204-206 °C; H NMR (500 MHz, Chloroform-d) δ 8.63 (d, J =
5.0 Hz, 2H), 8.24 (s, 1H), 7.86 (s, 1H), 7.69 (d, J = 5.1 Hz, 2H), 7.22 (d, J = 8.4 Hz, 1H), 7.10 (d,
J = 8.4 Hz, 1H), 5.32 (s, 2H), 3.60 (s, 3H), 2.87 (hept, J = 7.1 Hz, 1H), 2.62 (hept, J = 7.3 Hz, 2H),
2.25 (d, J = 12.7 Hz, 1H), 2.12 (d, J = 12.2 Hz, 1H), 1.79 – 1.65 (m, 4H), 1.65 – 1.59 (m, 1H),
1.40 (qd, J = 13.8, 13.1, 5.0 Hz, 2H), 1.22 (s, 3H), 1.15 (s, 3H), 1.10 (d, J = 6.9 Hz, 3H), 1.07 (d, J
= 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.1, 163.9, 150.4, 148.2, 145.8, 142.8, 137.5,
132.5, 130.5, 124.9, 123.5, 123.0, 120.0, 53.2, 52.0, 47.5, 44.1, 38.0, 37.1, 37.1, 36.5, 28.5, 28.4,
26.2, 25.1, 23.6, 23.2, 21.2, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C30H38N5O3,
516.2975, found, 516.2985; Anal. Calc. for C30H37N5O3: C, 69.88; H, 7.23; N, 13.58; O, 9.31.
Found: C, 70.01; H, 7.35; N, 13.67; O, 9.42.
4.2.7.25
14-(2-(4-((4,6-dichloro-1,3,5-triazin-2-ylamino)methyl)-1H-1,2,3-triazol-1-
yl)acetamido)dehydroabietic acid methyl ester(33)
White solid, (30.8 mg, 50%), mp 193-195 °C; 1H NMR (500 MHz, Chloroform-d) δ 7.96 (s, 1H),
7.70 (s, 1H), 7.49 (s, 1H), 7.22 (d, J = 8.3 Hz, 1H), 7.09 (d, J = 8.3 Hz, 1H), 5.24 (s, 2H), 4.74 (s,
2H), 3.62 (s, 3H), 2.83 (p, J = 6.9 Hz, 1H), 2.58 (d, J = 7.9 Hz, 2H), 2.25 (d, J = 12.9 Hz, 1H),
2.10 (d, J = 12.1 Hz, 1H), 1.68 (dd, J = 28.4, 15.8 Hz, 5H), 1.38 (q, J = 10.6, 7.4 Hz, 2H), 1.23 (s,
3H), 1.15 (s, 3H), 1.09 (d, J = 6.6 Hz, 3H), 1.05 (d, J = 6.7 Hz, 3H); 13C NMR (125 MHz, CDCl3)
δ 179.4, 170.9, 170.1, 165.7, 164.2, 148.2, 142.8, 132.5, 130.4, 124.9, 124.4, 123.5, 53.2, 52.1,
47.5, 44.1, 38.0, 37.1, 36.6, 28.4, 26.2, 25.1, 23.6, 23.2, 21.2, 18.5, 16.4; HRMS-ESI (m/z) [M +
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H]+ calcd for C29H37Cl2N8O3, 615.2366, found, 615.2369; Anal. Calc. for C29H36Cl2N8O3: C,
56.58; H, 5.89; N, 18.20; O, 7.80. Found: C, 56.69; H, 5.95; N, 18.23; O, 7.90.
4.2.7.26 14-(2-(4-Thiophen-2-yl)-1H-1,2,3-triazol-1-yl)acetamido)dehydroabietic acid methyl
ester(34)
White solid, (42.7 mg, 82%), mp 158-160 °C; 1H NMR (500 MHz, Chloroform-d) δ 7.96 (s, 1H),
7.44 (s, 1H), 7.39 (t, J = 4.0 Hz, 1H), 7.33 (d, J = 5.2 Hz, 1H), 7.23 (d, J = 8.4 Hz, 1H), 7.12 –
7.08 (m, 2H), 5.27 (s, 2H), 3.62 (s, 3H), 2.85 (hept, J = 6.8 Hz, 1H), 2.66 – 2.53 (m, 2H), 2.32 –
2.22 (m, 1H), 2.15 (d, J = 12.5 Hz, 1H), 1.72 (p, J = 11.1 Hz, 4H), 1.61 (d, J = 6.6 Hz, 1H), 1.44 (t,
J = 12.2 Hz, 1H), 1.37 (dd, J = 11.7, 5.2 Hz, 1H), 1.23 (s, 3H), 1.16 (s, 3H), 1.12 (d, J = 6.8 Hz,
3H), 1.09 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ 179.0, 164.0, 148.2, 142.8, 132.5,
132.0, 130.4, 127.8, 125.7, 124.9, 124.8, 123.4, 122.0, 53.4, 52.0, 47.5, 44.0, 38.0, 37.1, 36.5, 28.4,
26.2, 25.1, 23.6, 23.2, 21.2, 18.5, 16.5; HRMS-ESI (m/z) [M + H]+ calcd for C29H37N4O3S,
521.2586, found, 521.2597; Anal. Calc. for C29H36N4O3S: C, 66.89; H, 6.97; N, 10.76; O, 9.22.
Found: C, 66.97; H, 7.08; N, 10.83; O, 9.34.
Acknowledgements
This work was partially supported by the National Natural Science Foundation of NFSC, China
(Number: 21502114), China Postdoctoral Science Foundation, China (Number: 2015M581677)
and Open Foundation of the State Key Laboratory of Drug Research, China (Number:
SIMM1601KF-05).
Supplementary data
Supplementary data related to this article can be found at http:XXXXXXXXXXX
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broad spectrum potent in vitro antitumor activities
effectively against drug resistance cancer cells
stable in liver microsomes
induction of apoptosis in cancer cells
low toxicity