Optimization of 7-alkene-3-quinolinecarbonitriles as Src kinase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2010.03.025

The 7-alkene-3-quinolinecarbonitrile 20, a potent inhibitor of Src enzymatic and cellular activity with IC₅₀ values of 2.1 and 58 nM, respectively, had comparable efficacy to bosutinib in a colon tumor xenograft study.

Full text of DOI:10.1016/j.bmcl.2010.03.025

Bioorganic & Medicinal Chemistry Letters 20 (2010) 2924–2927
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Optimization of 7-alkene-3-quinolinecarbonitriles as Src kinase inhibitors
a
a
a
a
a
Diane H. Boschelli a, , Daniel Wang , Yan Wang , Biqi Wu , Erick E. Honores , Ana Carolina Barrios Sosa ,
*
Inder Chaudhary b, Jennifer Golas c, Judy Lucas c, Frank Boschelli c
a Wyeth Research, Chemical Sciences 401 N. Middletown Road, Pearl River, NY 10965, United States
b Wyeth Research, Drug Safety and Metabolism 401 N. Middletown Road, Pearl River, NY 10965, United States
c Wyeth Research, Oncology 401 N. Middletown Road, Pearl River, NY 10965, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
The 7-alkene-3-quinolinecarbonitrile 20, a potent inhibitor of Src enzymatic and cellular activity with
IC50 values of 2.1 and 58 nM, respectively, had comparable efficacy to bosutinib in a colon tumor xeno-
graft study.
Received 25 January 2010
Revised 27 February 2010
Accepted 4 March 2010
Available online 7 March 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
3-Quinolinecarbonitrile
Src
Kinase
Next year will mark the 100th anniversary of Peyton Rous’s dis-
covery that a virus can act as a transmissible agent and induce sar-
comas in healthy animals.1 It was subsequently determined that a
single gene contained in this virus was responsible for the tumor
formation.2 Additional studies established that this gene, which
was the first known oncogene, encoded a tyrosine kinase, v-Src,
which has a normal cellular counterpart, c-Src (Src). Activation of
Src causes aberrant cell signaling that can contribute to cancer.
Over the years considerable efforts have been expended to identify
small molecule inhibitors of Src kinase activity as potential anti-
cancer therapeutics.3–5 Three ATP competitive Src inhibitors are
currently in clinical trials for the treatment of solid tumors6: dasat-
inib (BMS-354825),7,8 saracatinib (AZD-0530),9,10 and bosutinib
(SKI-606).11,12
We previously reported that replacement of the C-7 alkoxy sub-
stituent on the 3-quinolinecarbonitrile core of bosutinib with cer-
tain alkenylaryl, 2a, or alkynylaryl, 2b, groups preserved the Src
kinase inhibitory activity.13,14 We also demonstrated that attach-
ment of a water solubilizing (dimethylamino)methyl group to the
aryl ring of the alkynylaryl analogs, as in 2c, provided potent Src
kinase inhibitors.15
Cl
Cl
O
HN
O
O
CN
N
N
N
1: Bosutinib
Cl
Cl
Cl
O
Cl
HN
HN
O
O
CN
O
CN
N
N
X
R
2a: X = N or CH
X
2b: X = N or CH; R = H
2c: X = N; R = CH2NMe2
second palladium catalyzed coupling reaction, now between 3a–c
and the 7-triflate-3-quinolinecarbonitrile 4,16 provided 5a–c. Con-
version of the primary alcohol group of 5a–c to a mesylate group
and subsequent displacement of the mesylate with dimethylamine
led to the desired 7-alkene analogs 6a–c. As seen in Table 1, the
2,5- and 2,6-isomers 6b and 6c were more potent Src inhibitors
than the 2,4-isomer 6a.
The C-7 alkene analogs of 2c where X is CH were prepared as
shown in Scheme 2. Palladium catalyzed coupling of 4-vinylbenz-
aldehyde17 and 3-vinylbenzaldehyde with triflate 4, provided 7a
and 7b, respectively. Subsequent reductive amination with
dimethylamine led to the desired 7-alkene analogs 8a and 8b.
The corresponding C-7 alkene analogs of 2c where X is N were
prepared as shown in Scheme 1. Palladium catalyzed coupling of 2-
bromo-4-(hydroxymethyl)pyridine with tributyl(vinyl)tin pro-
vided intermediate 3a. The 2,5- and 2,6-isomers, 3b–c, were pre-
pared in a similar fashion from the isomeric starting materials. A
* Corresponding author. Tel.: +1 845 602 3567; fax: +1 845 602 5561.
E-mail addresses: bosched@wyeth.com, dboschelli@optonline.net (D.H. Boschelli).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.03.025

D. H. Boschelli et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2924–2927
2925
Br
assay were evaluated for their 24 h plasma levels in nude mice
after a single oral 50 mg/kg dose. As shown in Table 1, none of
these three compounds had greater plasma levels than those of
bosutinib. When the two compounds with the highest plasma lev-
els, 6b and 6c, were tested in an HT-29 xenograft model with oral
dosing at 150 mg/kg, qd, for 14 days, neither compound was effica-
cious in blocking tumor growth.
We next replaced the C-7 alkenylaryl group with an alkenylalk-
yl group. As shown in Scheme 3, the initial analogs were prepared
from known vinyltributylstannanes, with the reaction of triflate 4
with 4-[(E)-3-(tributylstannyl)prop-2-enyl]morpholine18 and 4-
[(E)-4-(tributylstannyl)but-3-enyl]morpholine19 providing 9a and
9b, respectively. The additional reagents with extended alkyl
chains were prepared from the respective alkynes via a route anal-
ogous to that in the literature. Subsequent Stille coupling of these
vinyltributylstannanes with triflate 4 provided 9c–e.
4
a
N
N
5
HO
HO
6
3a: 2,4-isomer
3b: 2,5-isomer
3c: 2,6-isomer
Cl
Cl
O
Cl
Cl
O
HN
HN
b
O
CN
O
CN
4
5
N
TfO
N
N
OH
5a: 2,4-isomer
5b: 2,5-isomer
5c: 2,6-isomer
4
6
c
Cl
Cl
O
HN
O
CN
As shown in Table 2, the best inhibitors of Src enzymatic activ-
ity were 9b and 9c, having IC50 values of 4.8 and 5.4 nM respec-
tively. Extending the alkyl chain caused a substantial decrease in
activity. The best inhibition of Src cell activity was seen with 9b,
which had an IC50 value of 180 nM. As shown in Scheme 3, the
des C-6 methoxy analog of 9b, namely 12, was prepared by
4
N
N
5
N
6a: 2,4-isomer
6b: 2,5-isomer
6c: 2,6-isomer
6
Scheme 1. Reagents: (a) Bu3Sn-CH@CH2, Pd(PPh3)2Cl2, CuI, toluene; (b) 3a–c,
Pd(OAc)2, P(o-tol)3, DMF, Et3N; (c) (1) MsCl, Et3N, THF, DMF; (2) Me2NH, THF.
reaction
of
4-[(E)-4-(tributylstannyl)but-3-enyl]morpholine
with the 7-bromo-3-quinolinecarbonitrile 10.20 Reduced activity
in both the Src enzyme and cell assays was observed with 12. Fur-
ther loss of activity was observed with the analog of 12 lacking the
Table 1
Src enzyme and cell inhibitory activity and nude mouse 24 h plasma levels of 7-
alkenylaryl-3-quinolinecarbonitriles
Cl
Cl
Cl
O
Cl
Cl
Cl
HN
O
O
CN
HN
HN
O
a
O
CN
O
CN
O
N
N
X
N
TfO
N
(CH2)n
N
9a: n = 1
9b: n = 2
9c: n = 3
9d: n = 4
9e: n = 9
4
Compound
number
X
Isomer
Src enzyme
IC50 (nM)22
Src cell
24 h Plasma
level (ng/mL)
IC50 (nM)24
Bosutinib
3.8
6.8
2.2
2.2
2.4
3.0
100
280
58
28
40
40
Cl
Cl
R1
6a
6b
6c
8a
8b
N
N
N
CH
CH
2,4
2,5
2,6
1,4
1,3
Cl
Cl
36
28
10
HN
N
HN
R1
a
CN
CN
100
N
Br
N
O
10: R1 = OMe
11: R1 = H
12: R1 = OMe
13: R1 = H
Cl
Cl
Cl
Cl
HN
O
HN
O
Scheme 3. Reagents: (a) Bu3Sn-CH@CH(CH2)n-morpholine, Pd(PPh3)4, NMP.
a
O
CN
O
CN
N
TfO
N
OHC
Table 2
Src enzyme and cell inhibitory activity of 7-alkenylalkyl-3-quinolinecarbonitriles
7a: 1,4-isomer
7b: 1,3-isomer
4
Cl
Cl
Cl
Cl
b
HN
R1
HN
O
R2
CN
O
CN
O
N
N
(CH2)n
N
R1
R2
n
Src enzyme
IC50 (nM)22
Src cell
N
Compound
number
8a: 1,4-isomer
8b: 1,3-isomer
IC50 (nM)24
9a
9b
9c
9d
9e
12
13
OMe
OMe
OMe
OMe
OMe
OMe
H
OMe
OMe
OMe
OMe
OMe
H
1
2
3
4
9
2
2
8.7
4.8
5.4
18
130
15
260
180
350
Scheme 2. Reagents: (a) 4 and 3-vinylbenzaldehyde, Pd(OAc)2, P(o-tol)3, DMF,
Et3N; (b) Me2NH, THF, Na(OAc)3BH, CH2Cl2, DMF, AcOH.
290
3200
1100
2800
As shown in Table 1, while both 8a and 8b were potent Src
inhibitors in the enzyme assay, 8a was 2.5-fold more potent than
8b in the Src cell assay. The three most potent compounds in this
H
41

2926
D. H. Boschelli et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2924–2927
Cl
Cl
O
5-methoxy group on the aniline ring. Reaction of the 7-bromo-3-
quinolinecarbonitrile 1114 with 4-[(E)-4-(tributylstannyl)but-3-
enyl]morpholine provided 13 which had IC50 values for the inhibi-
tion of Src enzyme and Src kinase activity of only 41 nM and
HN
Sn(Bu)3
a
N
O
CN
N
2.8
lM, respectively.
N
N
The C-8 and C-6 isomers of 12, namely 15 and 18, were prepared
N
20
as shown in Scheme 4. Addition of 2,4-dichloro-5-methoxyaniline to
the 7-bromo-4-chloro-3-quinolinecarbonitrile 14 followed by reac-
tion with 4-[(E)-4-(tributylstannyl)but-3-enyl]morpholine pro-
vided the 8-alkenyl-3-quinolinecarbonitrile 15. The corresponding
C-6 isomer 18 was prepared from the 6-bromo-3-quinolinecarbo-
nitrile 16.20 As shown in Table 3, greatly reduced activity in the Src
enzyme assay was observed with both these isomers. Once again
further loss of activity was observed with 19, the analog of 18 lacking
the 5-methoxy group on the aniline ring.
The SAR to this point demonstrated the importance of the 5-
methoxy group on the aniline, the 6-methoxy on the 3-quinoline-
carbonitrile ring, and of the but-3-enyl group at C-7 of the core, in
agreement with what was observed in our earlier SAR studies on
related 3-quinolinecarbonitrile Src inhibitors. Since 9b had the best
Src enzyme and cell inhibitory activity, additional analogs were
prepared varying the water solubilizing amine group, starting with
the N-methylpiperazine 20.
As shown in Scheme 5, reaction of triflate 4 with 1-methyl-4-
[(3E)-4-(tributylstannyl)but-3-enyl]piperazine21 provided the de-
sired compound. Replacement of the morpholine group of 9b with
an N-methylpiperazine increased the Src enzyme inhibitory activ-
ity twofold (IC50 = 2.1 nM) and the cell inhibitory activity threefold
(IC50 = 58 nM). In a nude mouse PK study, a 50 mg/kg single oral
dose of 20 provided a 24 h plasma level of 38 ng/mL. Bosutinib
and 20 were tested side-by-side in an HT-29 xenograft study via
oral administration at 150 mg/kg, qd, for 14 days. When the tu-
Cl
Cl
O
HN
O
CN
TfO
N
4
b
Cl
Cl
O
Cl
Cl
HN
N
HN
O
O
CN
O
CN
c
R1R2N
N
HO
22a: NR1R2 = dimethylamine
22b: NR1R2 = piperidine
21
22c: NR1R2 = N-Et-piperazine
Scheme 5. Reagents: (a) 4, Pd(PPh3)4, NMP; (b) Bu3SnCH@CH(CH2)2-OH, Pd(PPh3)4,
NMP; (c) (1) MsCl, Et3N, THF, DMF; (2) R1R2NH.
mors were measured on day 21, both compounds were efficacious
with bosutinib having a T/C of 0.43 and 20 having a T/C of 0.36.
Additional profiling of 20 revealed that like bosutinib22 this 7-
alkene analog was also an inhibitor of Abl kinase activity, with
20 having an IC50 value of 0.4 nM. Weak activity was observed
against EGFR (IC50 = 3.3
other kinases including IKK, AKT, TPL2, PDK1, IGFR and mTOR
(IC50s >10 M). In a rat liver microsome stability assay, 20 had a
lM) and 20 was inactive against several
l
half-life of only 10 min. A short half-life was also observed in hu-
man microsomes (11 min), while in nude mouse microsomes 20
had a half-life of 33 min. Metabolite ID studies showed no metab-
olism occurred on the alkene group and that the major metabolites
included oxidation and dealkylation of the N-methylpiperazine.
To determine if replacement of the N-methylpiperazine group
of 20 with other amines would increase the microsomal stability,
additional analogs were targeted. As shown in Scheme 5, coupling
of triflate 4 with (E)-4-(tributylstannyl)-3-buten-1-ol23 provided
21, which contains a terminal alcohol group. Reaction of 21 with
mesyl chloride followed by treatment of the intermediate mesylate
with various amines gave the desired analogs 22a–c. As shown in
Table 4, all three compounds had similar Src inhibitory activity
to that of 20. In addition, all three had half-lives of greater than
30 min in a rat liver microsome stability assay.
Cl
Cl
O
Cl
N
HN
CN
CN
a,b
N
Br
O
14
15
N
Cl
Cl
Cl
Cl
R1
O
HN
R1
HN
N
CN
Br
CN
b
N
N
18: R1 = OMe
19: R1 = H
16: R1 = OMe
17: R1 = H
Scheme 4. Reagents: (a) 2,4-dichloro-5-methoxyaniline, 2-ethoxyethanol; (b)
Bu3Sn-CH@CH(CH2)2-morpholine, Pd(PPh3)4, NMP.
Table 4
Table 3
Src enzyme and cell inhibitory activity of 7-(but-3-enyl)-3-quinolinecarbonitriles
Src enzyme inhibitory activity of C-7, C-8 and C-6 substituted 4-(morpholin-4-yl)but-
1-enyl-3-quinolinecarbonitriles
Cl
Cl
Cl
Cl
HN
O
O
CN
HN
R1
O
R1R2N
N
CN
N
Compound
number
R1R2N
Src enzyme
IC50 (nM)22
Src cell
N
IC50 (nM)24
Compound number
Isomer
R1
Src enzyme IC50 (nM)22
15
9b
20
22a
22b
22c
Morpholine
N-Me-Piperazine
Me2N
Piperidine
N-Et-Piperazine
4.8
2.1
2.5
2.6
1.7
180
58
62
78
67
12
15
18
19
7
8
6
6
OMe
OMe
OMe
H
52% at 10
340
lM
1200

D. H. Boschelli et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2924–2927
2927
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Powell, D. W.; Yaczko, D.; Young, M.; Tischler, M.; Arndt, K.; Discafani, C.;
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While the Stille reactions of 4 with the vinyltributylstannanes
provided the desired products as mixtures of the E and Z isomers
in a ratio of approximately 98:2, Suzuki reaction of 4 with an
(E)-vinylboronic ester, namely 1-methyl-4-[(3E)-4-(4,4,5,5-tetra-
methyl-1,3,2-dioxaborolan-2-yl)-3-buten-1-yl]piperazine, provided
20 as >99% of the trans isomer.21 The identification of this highly
stereoselective route to the E alkenes, coupled with the in vivo effi-
cacy of 20 and the increased microsomal stability of 22a–c provides
rationale for further study of these 7-alkene-3-quinolinecarbonitrile
Src inhibitors.
12. Boschelli, D. H.; Boschelli, F. Drugs Future 2007, 32, 481.
13. Barrios Sosa, A. C.; Boschelli, D. H.; Wu, B.; Wang, Y.; Golas, J. M. Bioorg. Med.
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Acknowledgments
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Chem. Lett. 2004, 14, 2155.
15. Wu, B.; Barrios Sosa, A. C.; Boschelli, D. H.; Boschelli, F.; Honores, E. E.; Golas, J.
M.; Powell, D. W.; Wang, Y. D. Bioorg. Med. Chem. Lett. 2006, 16, 3993.
16. Berger, D.; Dutia, M.; Powell, D.; Wissner, A.; DeMorin, F.; Raifeld, Y.; Weber, J.;
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We thank the Wyeth Chemical Technology department for
compound characterization and the rat liver microsome stability
assays, Lin Deng and Ala Nassar of the Wyeth Drug Safety and
Metabolism department for the extensive metabolism studies of
20, Antonia Nikitenko of the Wyeth Discovery Synthetic Chemistry
department for the preparation of multi gram batches of triflate 4
and Tarek Mansour for his support.
References and notes
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