Synthesis and biological evaluation of a series of novel salicylanilides as inhibitors of EGFR protein tyrosine kinases

Article abstract of DOI:10.1016/j.cclet.2012.03.016

The synthesis and biological evaluation of two series of salicylanilide derivatives on the EGFR and ErbB-2 tyrosine kinases inhibitory activities were conducted. Of the tested compounds those having an additional aryl group substituted on the anilino ring were active on the EGFR tyrosine kinase inhibition (7a-c and 13a, 13c, 13d, 13f). The inhibitory activities were all in the low micromolar or submicromolar range. In addition, compound 13a was found to have dual inhibitory activities both on EGFR and ErbB-2 tyrosine kinases (1.654 ± 1.280 and 7.134 ± 1.265 μmol/L).

Full text of DOI:10.1016/j.cclet.2012.03.016

Available online at www.sciencedirect.com
Chinese Chemical Letters 23 (2012) 529–532
www.elsevier.com/locate/cclet
Synthesis and biological evaluation of a series of novel
salicylanilides as inhibitors of EGFR protein tyrosine kinases
Ning Ding a, Wei Zhang a, Hua Ling Xiao b, Peng Wang b, Ying Xia Li a,
*
a Department of Medicinal Chemistry, School of Pharmacy, Fudan University, Shanghai 201203, China
b School of Medicine and Pharmacy, Ocean University of China, Qingdao 266003, China
Received 2 December 2011
Available online 18 April 2012
Abstract
The synthesis and biological evaluation of two series of salicylanilide derivatives on the EGFR and ErbB-2 tyrosine kinases
inhibitory activities were conducted. Of the tested compounds those having an additional aryl group substituted on the anilino ring
were active on the EGFR tyrosine kinase inhibition (7a–c and 13a, 13c, 13d, 13f). The inhibitory activities were all in the low
micromolar or submicromolar range. In addition, compound 13a was found to have dual inhibitory activities both on EGFR and
ErbB-2 tyrosine kinases (1.654 ꢀ 1.280 and 7.134 ꢀ 1.265 mmol/L).
# 2012 Ying Xia Li. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: Salicylanilides; EGFR; HER2; Tyrosine kinase inhibitor
ErbB family type I receptor tyrosine kinases (EGFR, HER2, ErbB-3 and ErbB-4) play a critical role in mediating
growth factor signaling [1]. Overexpression of EGFR and/or HER-2 (ErbB-2) has been associated with oncogenic
activity such as unregulated cell growth, proliferation, differentiation and survival [2]. For this reason, they become
attractive targets for the design of novel anticancer drugs [3–6]. Two main approaches, humanized monoclonal
antibodies and tyrosine kinase inhibitors, have been developed. The ability to disrupt the ErbB-family signaling
pathway either by monoclonal antibody therapy [7] or small molecule ATP-competitive kinase inhibitors has
yielded novel anticancer agents [8]. In the last few years, a large structural variety of compounds, such as
4-anilinoquinazolines [9–11], 4-anilinopyrazolo[3,4-d]pyrimidines [12], 4-anilinoquinoline-3-carbonitriles [13],
4-anilinopyrazolo- and 4-anilinopyrroloquinazolines [14], were reported as EGFR tyrosine kinase inhibitors. As we
know, Iressa (1) [15] and Tarceva (2) [16] (Fig. 1) have been developed and approved for the chemotherapeutic
treatment of patients with advanced non-small-cell-lung cancer. The dual EGFR/HER-2 inhibitor Lapatinib (3) [17]
was recently approved for the treatment of HER2-positive metastatic breast cancer [18]. The latter possesses a novel
anilino-aryl head-group that yields dual EGFR/ErbB-2 potency (Fig. 1).
These agents all belong to the 4-anilinoquinazoline class of inhibitors and the key features between the receptor and
this template have been revealed as follows [19,20]. The quinazoline moiety fits into the ATP binding pocket of the
kinase domain, where the N-1 nitrogen of the quinazoline nucleus interacts with the backbone NH of Met-769 via a
* Corresponding author.
E-mail address: liyx417@fudan.edu.cn (Y.X. Li).
1001-8417/$ – see front matter # 2012 Ying Xia Li. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
doi:10.1016/j.cclet.2012.03.016

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N. Ding et al. / Chinese Chemical Letters 23 (2012) 529–532
O
F
F
O
S
H
N
Cl
Cl
HN
N
HN
N
HN
N
O
O
O
O
O
O
N
N
N
O
O
O
N
Tarceva (2)
Lapatinib (3)
Iressa (1)
Fig. 1. Structures of Iressa, Tarceva and Lapatinib.
hydrogen bond, and water mediated hydrogen bonding is observed between the N-3 of the quinazoline and the Thr-766
side chain. In addition, the aniline ring fills an adjacent lipophilic pocket. Furthermore, the solubilizing side chains at
C-6 and/or C-7 of the quinazoline core act to improve physical properties and confer a more favorable
pharmacokinetic profile. Many more compounds of this type are still under evaluation in clinical trials for the
treatment of cancer [21].
According to the results of Hodge and Traxler that revealed a bioisosteric relationship between the salicylic group
and the quinazoline (illustrated in Fig. 2) [22,23], the pseudo six-membered ring formed by the intramolecular
hydrogen bridge between OH and the carbonyl oxygen would function as a mimic of the pyrimidine ring of the
quinazoline [24]. The biological activities of salicylanilides are usually positively related to the electron-withdrawing
ability of the substituent at the anilino moiety [25], because the substituent of higher electron-withdrawing ability
facilitates the closed-ring conformation of salicylanilides required to be functional [26]. So far many salicylanilides
derivatives were synthesized using salicylic acid core structure function as an important pharmacophore group [25,27].
Enlightened by the recently reported small molecule EGFR-tyrosine kinases inhibitor such as Tarceva (2) and
Lapatinib (3), we would like to report here the design, synthesis and biological activity of a series of salicylanilide
derivatives as new tyrosine kinase inhibitors which specifically target EGFR and/or ErbB-2 tyrosine kinases. We
reserved the solubilizing side chains of 2 and 3 and put them into the salicylic acid core structure to construct two series
of salicylanilides derivatives 7a–c and 13a–h (Scheme 1), respectively. At the same time, different groups were
introduced to 30- and/or 40-positions of the anilino portion to see if they could keep or enhance the interaction with the
EGFR and/or HER-2 tyrosine kinase.
The synthetic routes towards the designed two series compounds are depicted in Scheme 1. Compounds 7a–c were
prepared from 4 [28] in three steps. Compound 4 was converted to 5 by treated with 10% NaOH aqueous solution and
then the amino-group of 5 was transformed into hydroxyl group according to the methodology reported by Cohen et al.
[29] to afford compound 6. Acylations of various of anilines by 2-hydroxy-4,5-bis(2-methoxy-ethoxy)benzoic acid 6
were achieved in the presence of EDCꢁHCl and HOAt (1-Hydroxy-7-azabenzotriazole) in DMF/THF (1:8) co-solvent
and afforded the corresponding target compounds 7a–c in good yield, respectively.
The synthetic route towards 13a–m is also depicted in Scheme 1. The first step was the coupling of commercially
available 2-hydroxy-5-iodobenzoic acid 8 with various anilines in the presence of EDCꢁHCl and HOAt in DMF/THF
(1:8) co-solvent to provide amides 9a–m. Then the palladium-catalyzed Suzuki coupling reaction [30] was adopted to
introduce the 5-formylfuran-2-yl into the amides 9a–m and to afford 11a–m. Finally, reductive amination of the
aldehyde with 2-aminoethylmethylsulfone 12 yielded the desired analogs 13a–m.
Target compounds 7a–c and 13a–m were evaluated for their EGFR and ErbB-2 tyrosine kinase activity following a
method described by Brignola and co-workers [31]. The IC50 values are listed in Table 1. It is interesting that of the
4'
3'
O
4'
3'
HN
O
HN
N
5
4
5
4
N
H
O
H
N
O
H
closed-ring
open-ring
salicylanilide
quinazoline
Fig. 2. Salicylanilide and quinazoline.

N. Ding et al. / Chinese Chemical Letters 23 (2012) 529–532
531
O
O
R =
O
O
O
O
O
OEt
(i)
O
O
OH
O
Cl
O
N
F
F
N
NH2
O
NH2
N
N
H
5
N
H
Cl
4
N
H
O
O
O
O
c
a
b
O
O
O
O
OH
(ii)
(iii)
O
O
R
OH
OH
O
O
O
6
7a-c
F
F
F
N
H
OMe
Me
N
N
H
H
f
d
OH
R
e
I
I
F
OH
(iv)
O
O
S
+
O
B
OHC
O
F
OH
OH
OH
N
H
N
H
Cl
8
10
9a-m
N
H
N
H
R
R
g
j
i
h
l
NH2
O
O
(v)
O
12
O
OH
OH
O
O
(vi)
O
HN
N
H
Cl
N
H
N
H
H
11a-m
13a-m
S
O
k
m
O
Scheme 1. Reagents and conditions: (i) 10% NaOH, 92%; (ii) NaNO2, concentrated H2SO4, 120 8C, 43%; (iii) RNH2, EDCꢁHCl, HOAt, DMF, 59%
for 7a, 49% for 7b, 67% for 7c; (iv) RNH2, EDCꢁHCl, HOAt, DMF/THF, 95% for 9a, 50% for 9b, 79% for 9c, 79% for 9d, 83% for 9e, 80% for 9f,
50% for 9g, 94% for 9h, 94% for 9i, 96% for 9j, 85% for 9k, 94% for 9l, 94% for 9m; (v) KOH, 5 mol% Pd(OAc)2, acetone/H2O, 35 8C, 66% for 11a,
29% 11b, 73% for 11c, 86% for 11d, 54% for 11e, 94% for 11f, 73% for 11g, 39% for 11i, 31% for 11j, 71% for 11k, 59% for 11l, 85% for 11m; (vi)
MgSO4, NaBH4, dry THF, reflux, 70% for 13a, 19% for 13b, 55% for 13c, 19% for 13d, 29% for 13e, 49% for 13f, 36% for 13g, 35% for 13h, 30%
for 13i, 29% for 13j, 33% for 13k, 36% for 13l, 38% for 13m.
Table 1
EGFR and ErbB-2 tyrosine kinases inhibition activity.a
Compound
EGFR
ErbB-2
Compound
EGFR
ErbB-2
IC50 (mmol/L)
IC50 (mmol/L)
IC50 (mmol/L)
IC50 (mmol/L)
7a
7b
0.780 ꢀ 0.380
0.990 ꢀ 0.400
0.300 ꢀ 0.218
1.654 ꢀ 1.280
>10
>10
>10
13f
13g
13h
13i
4.928 ꢀ 5.366
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
7c
>10
13a
13b
13c
7.134 ꢀ 1.265
>10
>10
13j
13k
13l
0.740 ꢀ 0.380
2.522 ꢀ 0.783
>10
13d
13e
>10
>10
13m
Lapatinib
0.006 ꢀ 0.001
0.044 ꢀ 0.016
a
IC50 values are shown as the mean ꢀ SD from three separate experiments.
tested compounds only those having an additional aryl group substituted on the anilino ring were active on the EGFR
tyrosine kinase inhibition (7a–c and 13a, 13c, 13d, 13f). The inhibitory activities were all in the low micromolar or
submicromolar range. In contrast, compounds 13f and g with no substituted aniline or a simply substituted anilino
moiety had no inhibitory activities on EGFR tyrosine kinase. In addition, of the active compounds 7a–c with two side
chains on the salicylic acid core structure were shown to be more active than those bearing a 5-
methylsulfonylethylamino-2-furanyl group (13a, 13c, 13d, 13f) generally. However, only compound 13a has dual
inhibitory activities both on EGFR and ErbB-2 tyrosine kinases (1.654 ꢀ 1.280 and 7.134 ꢀ 1.265 mmol/L).
Although the activity of 13a is lower than Lapatinib, it can be a start lead compound for these types of compounds at
the very beginning and could be further optimized.
In summary, we have conducted the synthesis and biological evaluation of two series of salicylanilide derivatives on
the EGFR and ErbB-2 tyrosine kinases inhibitory activities. The preliminary structure–activity relationships were
discussed and several compounds were found to be potent inhibitors of EGFR protein tyrosine kinases. All the results

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N. Ding et al. / Chinese Chemical Letters 23 (2012) 529–532
obtained here will render new clues to the understanding of the tyrosine kinases inhibitory profiles for these types of
compounds.
Acknowledgments
This work is supported by the Research Fund for the Doctoral Program of Higher Education of China (No.
20100071120051) and the Fundamental Research Funds for the Central Universities (No. 10FX061).
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