Design and Optimization of 3′-(Imidazo[1,2- a]pyrazin-3-yl)-[1,1′-biphenyl]-3-carboxamides as Selective DDR1 Inhibitors

Article abstract of DOI:10.1021/acsmedchemlett.9b00495

DDR1 is considered as a promising target for cancer therapy, and selective inhibitors against DDR1 over other kinases may be considered as promising therapeutic agents. Herein, we have identified a series of 3′-(imidazo[1,2-a]pyrazin-3-yl)-[1,1′-biphenyl]

Full text of DOI:10.1021/acsmedchemlett.9b00495

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Letter
Design and Optimization of 3'-(imidazo[1,2-a]pyrazin-3-yl)-
[1,1'-biphenyl]-3-carboxamides as Selective DDR1 Inhibitors
Cheng Mo, Zhang Zhang, Yupeng Li, Minhao Huang, Jian Zou, Jinfeng
Luo, Zheng-Chao Tu, Yong Xu, Xiaomei Ren, Ke Ding, and Xiaoyun Lu
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.9b00495 • Publication Date (Web): 06 Jan 2020
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Page 1 of 7
ACS Medicinal Chemistry Letters
Design and Optimization of 3'-(imidazo[1,2-a]pyrazin-3-yl)-[1,1'-
biphenyl]-3-carboxamides as Selective DDR1 Inhibitors
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Cheng Mo†, ‡, #, Zhang Zhang†, #, Yupeng Li‡, , #, Minhao Huang‡, Jian Zou†, Jinfeng Luo‡, Zheng-Chao
Tu‡, Yong Xu ‡, Xiaomei Ren†,*, Ke Ding†,*, Xiaoyun Lu†,*
†
International Cooperative Laboratory of Traditional Chinese Medicine Modernization and Innovative Drug
Development, Ministry of Education (MOE) of PR China, College of Pharmacy, Jinan University, 601 Huangpu
Avenue West, Guangzhou 510632, China
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‡
Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, 190 Kaiyuan Avenue, Guangzhou
510530, China

Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, United States
KEYWORDS: DDR1, Selective inhibitor, NSCLC, Drug discovery, SAR.
ABSTRACT: DDR1 is considered as a promising target for cancer therapy and selective inhibitors against DDR1 over other
kinases may be considered as promising therapeutic agents. Herein, we have identified a series of 3'-(imidazo[1,2-
a]pyrazin-3-yl)-[1,1'-biphenyl]-3-carboxamides as novel selective DDR1 inhibitors. Among these, compound 8v potently
inhibited DDR1 with an IC50 of 23.8 nM, while showed less inhibitory activity against DDR2 (IC50 = 1740 nM) and
negligible activities against Bcr-Abl (IC50 > 10 µM) and c-Kit (IC50 > 10 µM). 8v also exhibited excellent selectivity in a
KINOMEscan screening platform with 468 kinases. This compound dose-dependently suppressed NSCLC cell
tumorigenicity, migration and invasion. Collectively, these studies support its potential application for treatment of
NSCLC.
N
CF3
Discoidin domain receptors (DDR1 and DDR2),
discovered by homology cloning in 1990s, are
H
N
N
O
N
O
N
O
N
CF3
N
1,2
O
N
N
N
transmembrane receptor tyrosine kinases (RTKs). Unlike
H
H
N
other RTKs, the endogenous ligands of DDRs are collagens.3,4
DDRs can undergo autophosphorylation to actively regulate
2 (DDR1-IN-1)
1 (7rh)
basic cellular processes by collagen binding,
e.g.
H
N
O
differentiation, adhesion, proliferation, survival and matrix
remodeling. 5-7 Earlier studies have indicated that DDRs may
serve as potential targets of treating some diseases, such as
fibrotic disorders, osteoarthritis, and cancers.8,9 For example,
DDR1 was widely implicated in cell survival and invasiveness
in a range of cancers, including lung cancer, hepatocellular
carcinoma and prostate cancer, etc.10,11 Overexpression or
mutation of DDRs have also been detected in many cancer
cell lines such as non-small cell lung cancer (NSCLC), breast
cancer and ovarian cancer.12-14 Experimental therapeutics
targeting DDR1 by siRNA have indicated its potential to
suppress tumorigenicity, inhibit lung cancer bone metastasis,
and enhance cancer cell chemosensitivity.15-17 Consequently,
DDR1 inhibitors could serve as new potential therapeutic
agents for cancer treatment.9
H
N
H2N
N
CF3
N
N
N
H
N
N
H
N
H
O
O
4
3
NH2
CF3
H
N
O
H
H
N
N
N
H
N
O
N
N
N
O
N
N
F
N
5
6
Figure 1. Structures of reported selective DDR1 inhibitors.
M704
N
N
N
N
N
N
Hinge
N
N
R4
M704
Scaffold
Hopping
N
R5
R6
Numerous multi-kinase inhibitors have been reported to
show good inhibitory activities against DDR1 and DDR2
functionality.9 However, few of them were developed
primarily by targeting DDRs. During recent five years, several
selective DDR1 inhibitors were reported with various
selectivity profiles (Figure 1).18-23 Compound 1 was the first
reported by our laboratory as a promising DDR1-selective
inhibitor for cancer therapy. However, its selectivity against
DDR1 over DDR2 (14 fold) and Abl (52 fold) is still relatively
inadequate.18 To further improve its selectivity profile, a
series of 3'-(imidazo[1,2-a]pyrazin-3-yl)-[1,1'-biphenyl]-3-
carboxamides were designed by featuring a novel substituted
phenyl linker instead of the alkyne group in compound 1 and
the nitrogen of imidazo[1,2-a]pyrazine forming a critical
hydrogen bond with Met704 of DDR1 (Figure 2).
New linker
R2
R1
Middle
O
O
HN
CF3
O
CF3
HN
HN
Tail
R3
N
N
N
N
8
7
1
Figure 2. Designing of new DDR1 inhibitors based on
compound 1.
The title compounds were readily synthesized as
described in Scheme 1. Substituted methyl 3'-bromo-[(1,1'-
biphenyl)]-3-carboxyl-ates 13 were prepared by coupling a
substituted
substituted methyl 3-iodobenzoates 10 or by coupling
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acid
9
with

ACS Medicinal Chemistry Letters
Page 2 of 7
substituted 1-bromo-3-iodobenzenes 11 with methyl 4-
ethyl-3-(4,4,5,5-tetra-methyl-1,3,2-dioxaborolan-2-
8.7 nM), although its selectivity against other homologous
kinases was unexpectedly decreased (Table 1).
1
2
3
4
5
6
7
8
yl)benzoate (12). Compound 13 reacted with
bis(pinacolato)diboron to give the intermediate 14, which
underwent the classical Suzuki coupling reaction to yield
the key substituted methyl 3'-(imidazo[1,2-a]pyrazin-3-
yl)-[1,1'-biphenyl]-3-carboxylates 15. The title compounds
As a new scaffold for DDR1 inhibitor, compound 8a
exhibited digital nM inhibitory activity against DDR1, and
served as
a promising lead molecule for further
optimization. Further computational docking studies
suggested that 8a bound to DDR1 with a classical type II
bind mode (Figure 3A). The imidazo[1,2-a]pyrazine
moiety of 8a formed a critical HB with the Met704 of
DDR1. The phenyl linker participated in an additional π-π
interaction with Phe785, and this contributes greatly to
its improved selectivity against Bcr-Abl and c-Kit. The
amide of 8a also formed two HBs with Glu672 and Asp784
of DDR1, respectively. The modeling results suggested
that the trifluoromethyl group of 8a was deeply bound in
8
were obtained through the amidation of the
intermediate 15 with different anilines under basic
conditions.
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Scheme 1. Synthesis of the designed new DDR1
inhibitors.
R5
O
I
O
+
a1
OH
Br
9a-9h
B
R1
R5
R2
R5
R2
10
9
OH
: R5 = H
: R5 = Me
R6
R4
R4
Br
R6
O
10a-10f, 10t-10w
: R1 = Et
a2
O
b
O
10g
10h
: R1 = Me
: R1 = i-Pr
a
DFG-out hydrophobic pocket and the 1-(4-
9k
B
O
O
O
methyl)piperazinyl moiety was extended to solvent region
of protein. So, these two parts were firstly eliminated to
investigate the contribution to DDR1 inhibitory activity,
respectively.
R1
14
R1
5'
R
13
O
B
O
6'
4'
+
O
O
Br
I
2'
11
: 2'-Me
: 4'-Me
12
11i
11j
11k
11p
11q
11r
: 5'-OMe
: 5'-CN
:5'-F
11s
11l
11n
: 5'-Cl
: 6'-Me
: 5'-i-Pr
: 5'-t-Bu
R5
R2
5'
2'
R3
R4
R6
6'
: 5'-Me
4'
11o
O
N
R5
N
3'
N
H
R4
R6
O
N
c
d
R1
: R3 = 3'-CF3, 5'-(4-methylpiperazin-1-yl)methyl
8b:
N
N
8
O
8a
R2
R1
N
R3 = 3'-(4-methylpiperazin-1-yl)methyl
8c 8g 8h 8i 8s
8t: R3 = 3'-CF3, 2'-F
,
,
,
-
: R3 = 3'-CF3
15
8u
: R3 = 3'-CF3, 4'-F
8d
8e
8f
: R3 = 4'-CF3
: R3 = 2'-CF3
: R3 = H
8v
: R3 = 3'-CF3, 5'-F
8w
: R3 = 3'-CF3, 6'-F
Reagents and conditions: (a1) Pd(PPh3)4, Na2CO3,
PhMe/H2O (3:1), Ar, 90 °C, overnight,58-98%; (a2) Pd(PPh3)4,
K3PO4, dioxane, Ar, 90 °C, overnight,55-91%; (b) Pd(dffp)Cl2,
AcOK, Bis(pinacolato)diboron, dioxane, Ar, 90 °C,
overnight,64-92%; (c) Pd(PPh3)4, Na2CO3, PhMe/H2O (3:1),
Ar, 90 °C, overnight, 42-82%; (d) t-BuOK, anilines, dry THF, -
20 °C, 4-79%.
Figure 3. Binding modes of compounds 8a (A) and 8c (B)
to DDR1 Kinase (PDB: 4bkj). HBs and π-π interactions
with key amino acids are indicated by yellow dotted lines .
Compound 8b in which R was replaced by 1-benzyl-4-
methylpiperazine, showed a substantial potency loss
towards DDR1 (IC50 = 357.1 nM), while compound 8c
without the 1-(4-methyl)piperazinyl group was almost
equal potent with 8a and had significantly improved
selectivity over DDR2, Bcr-Abl and c-Kit (Table 1). The
results suggested the CF3 group is essential for the
compounds to maintain DDR1 inhibitory activity and
improve the selectivity over the aforementioned kinases.
Moving the CF3 group to the ortho- (8d) or para- (8e)
positions, or simply eliminating it (8f) totally negated the
inhibitory activity against DDR1 kinase. The contribution
of R1 substituents to the DDR1 potency and selectivity was
also investigated. The methyl-substituted compound (8g)
totally abolished the selectivity over DDR2, Bcr-Abl and c-
Kit (Table 1), although the inhibitory activity was
maintained. The isopropyl-substituted compound (8h)
also totally abolished the activity. These results clearly
suggested that R1 group, directed towards the gatekeeper
Thr701 (Figure 3), played a crucial role in regulating
kinase selectivity and activity.
Our previous study indicated that 1 bound to the
active pocket of DDR1 with a type II binding mode, and
the alkynyl moiety served as a linking group for the
pyrazolo[3,4-b]pyridine and the ethylbenzene.24 In our
continuous efforts to improve its selectivity, the
conformationally restricted bioisostere phenyl group of
the alkynyl was first explored (Figure 2), similar to that of
bioisostere triazol. 25
Starting with the simplest modification, compound 7
indeed improved the selectivity against Bcr-Abl and c-Kit,
although it had about 50-fold decreased inhibitory
activities against DDR1 and DDR2 (Table 1). In an
investigation of the reason for the loss of activity,
modeling studies suggested that compound 7 did not fit
nicely into the DDR1 ATP binding pocket and could not
achieve the essential hydrogen bond (HB) with Met704 in
DDR1 (Figure 2). In view of the importance of the HB
interactions in the hinge region of kinases, imidazo[1,2-
a]pyrazine was used as a privileged moiety to restore the
key HB formed with Met704 of DDR1.26 Compound 8a
Table 1. Kinase inhibition of compounds 7 and 8a-8h against
DDR1, DRR2, Bcl-Abl, and c-Kit.
exhibited 37-fold increased potency against DDR1 (IC50
=
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ACS Medicinal Chemistry Letters
and a detailed SAR study focusing on phenyl linker was
O
N
1
2
3
4
5
6
7
8
undertaken. When a methyl group was added at the R2,
R4, R5 or R6 position of 8c, the resulting compounds 8i-8l
showed reduced inhibitory activity against DDR1 with IC50
values of 30.1, 107.8, 22.5 and 63.7 nM, respectively.
Although 8k in which R5 as methyl, was about 2-fold less
potent than 8c, it displayed superior selectivity over other
three kinases (DDR2, Bcr-Abl and c-Kit). Next, we
examined the contribution of R5 substituent to the DDR1
inhibitory potency and selectivity. Obviously, as the bulk
of the substituents increased, the potency against DDR1
decreased accordingly. For example, replacement of the
R5-methyl with ethyl (8m), isopropyl (8n), or tert-butyl
(8o) led to compounds with IC50 values of 42.3, 66.9 and
92 nM, respectively. Further study showed that this
position was generally tolerant to both medium sized
electron-donating and -withdrawing groups. For example,
compounds with methoxy (8p), cyano (8q), fluorine (8r),
and chlorine (8s) groups at this position exhibited IC50
values of 16.5, 15.6, 30.3 and 21.3 nM, respectively, against
the DDR1 kinase, but the selectivity over DDR2, Bcr-Abl
and c-Kit was obviously decreased.
R
N
N
H
N
R1
Kinase inhibition (IC50, nM)
Cpds
R1
R
DDR1
DDR2
Bcr-
c-Kitb
a
a
Ablb
CF3
O
N
N
N
N
H
N
N
7
359.5
196.7
>10µM >10µM
9
CF3
N
10
11
12
13
14
15
16
17
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N
8a
8b
Et
Et
9.6
66.1
437
619.7
4934
N
N
357.1
>10µM
989.3
CF3
8c
8d
8e
8f
8g
8h
1
Et
Et
Et
Et
10.4
1296
4879
4850
9.2
725
2220
4992
2917
CF3
>10µM
3984
>10µM >10µM >10µM
CF3
>10µM
135
5120
237
8990
72.4
CF3
CF3
M
e
i-
The fluorine atom is a bioisostere of hydrogen and can
reduce the electrical properties of the phenyl group. We
investigated the potential influence of the “tail phenyl”
region of 8k by introducing a fluorine atom to the R
position. The resulting 2’-F (8t), 4’-F (8u), and 6’-F (8w)
compounds displayed decreased inhibitory activity
against DDR1, while 5’-F (8v) exhibited a similar DDR1
inhibitory activity and selectivity to those of 8k.
Compound 8v potently exhibited DDR1 inhibitory activity
(IC50 = 23.8 nM) and also targeted selectivity over
aforementioned kinases DDR2, Bcr-Abl and c-Kit of 73-,
420-, and 420-fold, respectively.
1260
6.8
>10µM >10µM
101.4 355
1686
Pr
-
>10µM
aDDR1 and DDR2 inhibition were performed with the
LanthaScreen Eu kinase assay. b Bcr-Abl and c-Kit inhibition
were performed with the FRET-based Z′-Lyte assay. *Data
means of three independent experiments, and the variations
are <20%.
Our previous study suggested that the π-π interaction
between the phenyl linker of 8c with Phe785 contributed
significantly to the selectivity against Bcr-Abl and c-Kit,23
Table 2. Kinase inhibition of compounds 8i−8y against DDR1, DRR2, Bcr-Abl, and c-Kit.
R5
5'
R
R4
R6
6'
O
4'
CF3
N
N
N
2'
H
R2
N
Kinase inhibition (IC50, nM)
Cpds
R2
R4
R5
R6
R
DDR1a
30.1
DDR2a
663.5
>10µM
1728
Ablb
c-Kitb
8i
8j
Me
H
H
H
H
H
H
H
H
H
H
Me
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
1813
5978
107.8
22.5
63.7
42.3
66.9
92
>10µM
>10µM
1452
>10µM
>10µM
>10µM
>10µM
>10µM
>10µM
4994
8k
8l
Me
H
H
H
Me
H
610
8m
8n
8o
8p
8q
8r
H
Et
3227
2631
>10µM
>10µM
>10µM
3696
H
i-Pr
t-Bu
OMe
CN
F
H
H
H
>10µM
1018
H
H
16.5
H
H
15.6
947
5637
3721
H
H
30.3
1326
5345
6591
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8s
8t
8u
8v
8w
1
H
H
H
H
H
H
H
H
H
H
Cl
Me
Me
Me
Me
-
H
H
H
H
H
H
21.3
73.8
55.6
23.8
36.5
6.8
1080
>5µM
>5µM
1740
>10µM
>10µM
>10µM
>10µM
>10µM
8414
1
2
3
4
5
6
7
8
2’-F
4’-F
5’-F
6’-F
>10µM
>10µM
>10µM
>10µM
>10µM
>5µM
101.4
355
b
aDDR1 and DDR2 inhibition were performed with the LanthaScreen Eu kinase assay. Bcr-Abl and c-Kit inhibition were
performed with the FRET-based Z′-Lyte assay. *Data means of three independent experiments, and the variations are <20%.
9
To further evaluate the selectivity of 8v over other
kinases, a KINOMEscan screening platform with 468 kinases
was conducted at DiscoveRx (San Diego, CA). 27 The results
demonstrated that 8v displayed extraordinary target
selectivity (S core (10) = 0.01 and S core (1) = 0.007) (Figure
4A, Tables S1 and S2). The apparent “off-target” kinases
included only DDR2, calcium dependent protein kinase ID
(CAMK1D), CDC like kinase 4 (CLK4), and spindle assembly
checkpoint kinase (TTK). The Kd values of 8v against above
“off targets” and DDR1 were further evaluated using
DiscoveRx’s platform. The results showed that 8v tightly
bound to DDR1 (Kd = 7.8 nM, Figure 4B), which was
consistent with the in vitro kinase inhibitory activity. For
potential “off-targets”, compound 8v had no detectable
binding affinities to three apparent “off target” kinases
(CAMK1D, CLK4, and TTK) at a >1 µM concentration except
for DDR2 (Figure 4B). Although Kd value of 8v to DDR2
appears to be as low as 86 nM, its IC50 value against DDR2
was 1739 nM in our kinase assay. In comparison, the IC50
value of compound 8v against DDR1 was determined to be
23.8 nM (73-fold selective). Taken together, these results
suggested the extraordinary target selectivity of compound
8v against a panel of 468 kinases.
significantly suppressed the number of colony formation
(not the size).
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Figure 5. Compound 8v suppresses H1299 NSCLC cell
tumorigenicity and proliferation. (A) Compound 8v
effectively inhibits H1299 NSCLC cell colony formation. (B)
The quantitative result of Figure 5A. (C) The proliferation
result of 8v evaluated with an CCK-8 assay. The data are
means from three independent experiments.
It has been reported that DDR1 plays important roles in
tumor migration and invasion.18 The effect of 8v on the
migration and invasion of H1299 NSCLC cancer cells was
investigated using the wound healing assay and the
transwell assay. As shown in Figure 6A, compound 8v
suppressed wound closure in H1299 cells by 40%, 60%
and 84% at concentrations of 1.25, 2.5 and 5.0 M,
respectively, compared with blank control groups.
Further the study showed that 8v dose-dependently
inhibits the invasion of H1299 cancer cells by 17%, 50%
and 71% at the 1.25, 2.5 and 5.0 M concentrations for 24
h, respectively (Figures 6B). These results clearly
indicated that compound 8v can potentially suppress
migration and invasion of H1299 NSCLC cancers.
Figure 4. KINOMEscan profiles of compound 8v. (A)
KINOMEscan profiling of 8v at a concentration of 200 nM
(25-fold Kd value) against 468 kinases. (B) The Kd
determination of compound 8v against DDR1 and the
potential off targets.
A previous study has shown that DDR1 inhibitors strongly
suppress cancer tumorigenicity.18 The effect of 8v on the
tumorigenicity of H1299 NSCLC cancer cells was firstly
investigated by colony formation assay. The results
demonstrated
that compound 8v dose-dependently
suppressed colony formation in H1299 NSCLC cancer cells
after 10 continuous day treatment of cells with 8v (IC50 =
0.046 µM, Figures 5A and 5B). However, the direct anti-
proliferation of 8v against H1299 NSCLC cancer cells
seemed to be moderate with an IC50 value of 6.6 µM
(Figure 5C). It was suggested that compound 8v
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ACKNOWLEDGMENT
1
2
3
4
This work was supported by National Natural Science
Foundation of China (81922062, 81874285 and 81820108029)
National Key Research and Development Program of China
(218YFE0105800) and Guangdong Province Science and
Technology Program (2018A050506043) and Jinan University.
5
6
7
ABBREVIATIONS
8
9
DDR1, discoidin domain receptor 1; Kd, binding constant;
RTKs, receptor tyrosine kinases; CF3, trifluoromethylphenyl;
CAMK1D, calcium/calmodulin dependent protein kinase ID;
NSCLC, non-small cell lung cancer; CLK4, CDC like kinase 4;
SAR, structure-activity relationship; TTK spindle assembly
checkpoint kinase; DFG, Asp-Phe-Gly; MTT, 3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenytetrazolium bromide.
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Figure 6. Compound 8v suppresses H1299 NSCLC cell
migration and invasion. (A) Compound 8v inhibition of
H1299 cell migration with wound healing assay. (B)
Compound 8v inhibition of H1299 cell migration and
invasion with transwell assay. The data are means from three
independent experiments.
REFERENCES
1. Johnson, J. D.; Edman, J. C.; Rutter, W. J. A receptor
tyrosine kinase found in breast carcinoma cells has an
extracellular discoidin I-like domain. Proc. Natl. Acad.
Sci. 1993, 90, 5677-5681.
2. Lai, C.; Lemke, G. Structure and expression of the Tyro
10 receptor tyrosine kinase. Oncogene, 1994, 9, 877-883.
3. Vogel, W.; Gish, G. D.; Alves, F.; Pawson, T. The
discoidin domain receptor tyrosine kinases are activated
by collagen. Mol. Cell. 1997, 1, 13-23.
4. Leitinger, B. Molecular analysis of collagen binding by
the human discoidin domain receptors, DDR1 and
DDR2. Identification of collagen binding sites in DDR2.
J. Biol. Chem. 2003, 278, 16761–16769.
5. Fu, H. L.; Valiathan, R. R.; Arkwright, R.; Sohail, A.;
Mihai, C.; Kumarasiri, M.; Mahasenan, K. V.; Mobashery,
S.; Huang, P.; Agarwal, G.; Fridman, R. Discoidin domain
receptors: unique receptor tyrosine kinases in collagen-
mediated signaling. J. Biol. Chem. 2013, 288, 7430-7437.
6. Hou, G.; Vogel. W.F.; Bendeck, M.P. Tyrosine kinase
activity of discoidin domain receptor 1 is necessary for
In summary, a series of 3'-(imidazo[1,2-a]pyrazin-3-yl)-
[1,1'-biphenyl]-3-carboxamides were synthesized and
investigated as highly selective DDR1 inhibitors. The
comprehensive SARs study reported here led to the most
promising compound 8v, which binds tightly to the DDR1
(Kd = 7.8 nM) and potently inhibits DDR1 kinase function
(IC50 = 23.8 nM). Furthermore, 8v exhibits ideal target
selectivity in KINOMEscan screen against the 468
kinases. Compound 8v also potently suppresses NSCLC
cell tumorigenicity, invasion and adhesion. However, the
pharmacokinetic parameters of compound 8v is not ideal
(t1/2 = 1.2 hr and F % = 3.9). Further pharmacokinetics-
oriented optimization of 8v is on-going and the results
will be disclosed in due course. Collectively, compound 8v
may be as a promising lead compound for the future
studies.
ASSOCIATED CONTENT
Supporting information
smooth
muscle
cell
migration
and
matrix
metalloproteinase expression. Circ. Res. 2002, 90, 1147-
1149.
Synthetic procedures for compounds 8a-8w, the results of
the kinase selectivity profiling study of compound 8v,
procedures for Kinomescan screening, in vitro kinase assay,
colony formation assay, in vitro cell proliferation by MTT
assay, wound healing, transwell assay, computational study
and the 1H and 13C NMR spectra of compounds 8a-8w will be
released upon article publication.
7. Vogel, W. F.; Abdulhussein, R.; Ford, C. E. Sensing
extracellular matrix: an update on discoidin domain
receptor function. Cell Signal. 2006, 18, 1108-1116.
8. Borza, C. M.; Pozzi, A. Discoidin domain receptors in
disease. Matrix Biol. 2014, 34, 185-192.
9. Li, Y.; Lu, X.; Ren, X.; Ding, K. Small molecule discoidin
domain receptor kinase inhibitors and potential medical
applications. J. Med. Chem. 2015, 58, 3287-3301.
10. Shimada, K.; Nakamura, M.; Ishida, E.; Higuchi, T.;
Yamamoto, H.; Tsujikawa, K.; Konishi, N. Prostate cancer
antigen-1 contributes to cell survival and invasion
through discoidin receptor 1 in human prostate cancer.
Cancer Sci. 2008, 99, 39-45.
11. Valiathan, R. R.; Marco, M.; Leitinger, B.; Kleer, C. G.;,
Fridman, R. Discoindin domain receptor tyrosine
kinases: new players in cancer progression. Cancer
Metastasis Rev. 2012, 31, 295-321.
The Supporting Information is available free of charge on the
ACS Publications website.
AUTHOR INFORMATION
Corresponding Author
*E-mail addresses: ren_xiaomei@jnu.edu.cn (X. R.), dingke
@jnu.edu.cn (K. D.), luxy2016@jnu.edu.cn (X. L.).
Author Contributions
# These authors contributed equally to this work.
Notes
12. Ford, C. E.; Lau, S. K.; Zhu, C. Q.; Andersson, T.; Tsao, M.
S.; Vogel, W. F. Expression and mutation analysis of the
discoidin domain receptors 1 and 2 in non-small cell lung
The authors declare no competing financial interest.
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ACS Medicinal Chemistry Letters
Page 6 of 7
carcinoma. Br. J Cancer. 2007, 96, 808-814.
Characterization of DDR2 inhibitors for the treatment of
DDR2 mutated non-small cell lung cancer. ACS Chem.
Biol. 2015, 10, 2687-2696.
1
2
3
4
5
6
7
8
13. Barker, K. T.; Martindale, J. E.; Mitchell, P. J.; Kamalati,
T.; Page, M. J.; Phippard, D. J.; Dale, T. C.; Gusterson, B.
A.; Crompton, M. R. Expression patterns of the novel
receptor-like tyrosine kinase, DDR, in human breast
tumors. Oncogene. 1995, 10, 569-575.
14. Quan, J.; Yahata, T.; Adachi, S.; Yoshihara, K.; Tanaka, K.
Identification of receptor tyrosine kinase, discoidin
domain receptor 1 (DDR1), as a potential biomarker for
serous ovarian cancer. Int. J Mol. Sci. 2011, 12, 971-982.
15. Valencia, K.; Ormazabal, C.; Zandueta, C.; Luis-Ravelo,
D.; Anton, I.; Pajares, M. J.; Agorreta, J.; Montuenga, L.
M.; Martínez-Canarias, S.; Leitinger, B.; Lecanda, F.
Inhibition of collagen receptor discoidin domain
receptor-1 (DDR1) reduces cell survival, homing and
colonization in lung cancer bone metastasis. Clin. Cancer
Res. 2012, 18, 969-980.
16. Das, S.; Ongusaha, P. P.; Yang, Y. S.; Park, J. M.;
Aaronson, S. A.; Lee, S. W. Discoidin domain receptor 1
receptor tyrosine kinase induces cyclooxygenase-2 and
promotes chemoresistance through nuclear factor-
kappaB pathway activation. Cancer Res. 2006, 66, 8123-
8130.
17. Kim, H. G.; Hwang, S. Y.; Aaronson, S. A.; Mandinova, A.;
Lee, S. W. DDR1 receptor tyrosine kinase promotes
prosurvival pathway through Notch1 activation. J. Biol.
Chem. 2011, 286, 17672-17681.
22. Murray, C. W.; Berdini, V.; Buck, I. M.; Carr, M. E.;
Cleasby, A.; Coyle, J. E.; Curry, J. E.; Day, J. E.; Day, P. J.;
Hearn, K.; Iqbal, A.; Lee, L. Y.; Martins, V.; Mortenson,
P. N.; Munck, J. M.; Page, L. W.; Patel, S.; Roomans, S.;
Smith, K.; Tamanini, E.; Saxty, G. Fragment-based
discovery of potent and selective DDR1/2 Inhibitors. ACS
Med. Chem. Lett. 2015, 6, 798-803.
23. Wang, Z.; Bian, H.; Bartual, S. G.; Du, W.; Luo, J.; Zhao,
H.; Zhang, S.; Mo, C.; Zhou, Y.; Xu, Y.; Tu, Z.; Ren, X.;
Lu, X.; Brekken, R. A.; Yao, L.; Bullock, A. N.; Su, J.;
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
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Ding,
K.
Structure-based
design
of
tetrahydroisoquinoline-7-carboxamides as selective
discoidin domain receptor 1 (DDR1) inhibitors. J. Med.
Chem. 2016, 59, 5911–5916.
24. Wang, Z.; Zhang, Y.; Pinkas, D. M.; Fox, A. E.; Luo, J.;
Huang, H.; Cui, S.; Xiang, Q.; Xu, T.; Xun, Q.; Zhu, D.;
Tu, Z.; Ren, X.; Brekken, R. A.; Bullock, A. N.; Liang, G.;
Ding, K.; Lu, X. Design, Synthesis, and Biological
Evaluation of 3-(Imidazo[1,2- a]pyrazin-3-ylethynyl)-4-
isopropyl-
N-(3-((4-methylpiperazin-1-yl)methyl)-5-
(trifluoromethyl)phenyl)benzamide as a Dual Inhibitor
of Discoidin Domain Receptors 1 and 2. J. Med. Chem.
2018, 61, 7977-7990.
25. Li, Y.; Shen, M.; Zhang, Z.; Luo, J.; Pan, X.; Lu, X.; Long,
H.; Wen, D.; Zhang, F.; Leng, F.; Li, Y.; Tu, Z.; Ren, X.;
Ding, K. J. Med. Chem. 2012, 55, 10033-10046.
26. Wang, Z.; Zhang, Y.; Bartual, S. G.; Luo, J. F.; Xu, T; Du,
W.; Xun, Q.; Tu, Z.; Brekken, R. A.; Ren, X.; Bullock, A.
N.; Liang, G.; Lu, X.; Ding, K. Tetrahydroisoquinoline-7-
carboxamide derivatives as new selective discoidin
domain receptor 1 (DDR1) Inhibitors. ACS Med. Chem.
Lett. 2017, 8, 327-332.
27. Fabian, M. A.; Biggs, W. H., 3rd; Treiber, D. K.;
Atteridge, C. E.; Azimioara, M. D.; Benedetti, M. G.;
Carter, T. A.; Ciceri, P.; Edeen, P. T.; Floyd, M.; Ford, J.
M.; Galvin, M.; Gerlach, J. L.; Grotzfeld, R. M.; Herrgard,
S.; Insko, D. E.; Insko, M. A.; Lai, A. G.; Lelias, J. M.;
Mehta, S. A.; Milanov, Z. V.; Velasco, A. M.; Wodicka, L.
M.; Patel, H. K.; Zarrinkar, P. P.; Lockhart, D. J. A small
molecule-kinase interaction map for clinical kinase
inhibitors. Nat. Biotechnol. 2005, 23, 329-336.
18. Gao, M.; Duan, L.; Luo, J.; Zhang, L.; Lu, X.; Zhang, Y.;
Zhang, Z.; Tu, Z.; Xu, Y.; Ren, X.; Ding, K. Discovery and
optimization
of
3-(2-(Pyrazolo[1,5-a]pyrimidin-6-
yl)ethyn yl)benzamides as novel selective and orally
bioavailable discoidin domain receptor
inhibitors. J. Med. Chem. 2013, 56, 3281−3295.
1
(DDR1)
19. Kim, H. G.; Tan, L.; Weisberg, E. L.; Liu, F.; Canning, P.;
Choi, H. G.; Ezell, S. A.; Wu, H.; Zhao, Z.; Wang, J.;
Mandinova, A.; Griffin, J. D.; Bullock, A. N.; Liu, Q.; Lee,
S. W.; Gray, N. S. Discovery of a potent and selective
DDR1 receptor tyrosine kinase inhibitor. ACS Chem. Biol.
2013, 8, 2145−2150.
20. Richters, A.; Nguyen, H. D.; Phan, T.; Simard, J. R.;
Grutter, C.; Engel, J.; Rauh, D. Identification of type II
and III DDR2 inhibitors. J. Med. Chem. 2014, 57, 4252-
4262.
21. Terai, H.; Tan, L.; Beauchamp, E. M.; Hatcher, J. M.; Liu,
Q.; Meyerson, M.; Gray, N. S.; Hammerman, P. S.
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