1
118 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 7
Letters
Sch em e 3. Synthesis of Amines for 12g and 12j
Ta ble 2. Effects of Protein Binding on Cellular Potencya
PDGF-Râ targets in biochemical and cellular assays,
solubility under both neutral and acidic conditions, and
protein binding properties. Additional kinase selectivity
study revealed that 12b is also a good inhibitor of KIT
PDGF-induced BrdU incorporation in 3T3 cells,
IC50 (µM)
at 0%
BSA
at 0.1%
BSA
at 0.5%
BSA
at
at
1
3
compd
1% BSA
5% BSA
and FLT-3. Furthermore, 12b also had very good oral
bioavailability, was highly efficacious in a number of
preclinical tumor models, and was well tolerated at
5
1
b
2b
0.3
<0.007
9.5
<0.007
16
<0.007
36
0.008
49
0.083
8
a
efficacious doses. It is currently in clinical phase I trials
BSA ) bovine serum albumin.
for the treatment of cancers.
inhibitory activity against VEGF-R2 and PDGF-Râ in
biochemical and cellular assays (Table 1). Specifically,
compared to 5b, 12b is about 30-fold more potent
against VEGF-R2 and PDGF-Râ in biochemical assays,
over 10-fold more potent in cellular kinase assays, and
significantly more soluble under neutral (20-fold) and
acidic (>500-fold) conditions. Among 12a -d , 12a is
markedly less potent in inhibiting PDGF-Râ phos-
phorylation in biochemical and cellular assays. Different
halogen substitutions had little effect on the biochemical
activities against VEGF-R2 and PDGF-Râ but affected
the cytotoxicity profiles. For example, the bulkier Cl and
Br substitutions (12c and 12d ) show some extent of
cytotoxicity when compared to the F analogue (12b)
Su p p or tin g In for m a tion Ava ila ble: Synthesis of 12b.
This material is available free of charge via the Internet at
http://pubs.acs.org.
Refer en ces
(
1) Cherrington, J . M.; Strawn, L. M.; Shawver, L. K. New para-
digms for the treatment of cancer; the role of anti-angiogenesis
agents. In Advances in Cancer Research, 1st ed.; Klein, G., Van
de Woude, G. F., Eds.; : Academic Press: San Diego, CA, 2000;
Vol. 70, pp 1-38.
(2) Gale, N. W.; Yancopoulos, G. D. Growth factors acting via endo-
thelial cell-specific receptor tyrosine kinases: VEGFs, angio-
poietins, and ephrins in vascular development. Genes Dev. 1999,
1
3, 1055-1066.
(3) Ferrara, N. VEGF: an update on biological and therapeutic
aspects. Curr. Opin. Biotechnol. 2000, 11, 617-624.
(Table 1). Therefore, diversification on 12b has been
(
4) Rosenkranz, S.; Kazlauskas, A. Evidence for distinct signaling
properties and biological responses induced by the PDGF recep-
tor alpha and beta subtypes. Growth Factors 1999, 16, 201-
further explored.
Surprisingly, replacing the diethylamine in 12b by
either dimethylamine (12e) or pyrrolidine (12f) mark-
edly reduced solubility at pH 6 (from 364 to 75 or 9 µg/
mL, respectively), while the in vitro activities were
similar (Table 1). The more basic N-methylpiperidine
group (which significantly increased solubility of the
quinazoline-based VEGF-R2/EGF-R inhibitor (4-bromo-
2
16.
(
5) Sun, L.; Tran, N.; Tang, T.; App, H.; Hirth, P.; McMahon, G.;
Tang, C. Synthesis and biological evaluations of 3-substituted
indolin-2-ones: a novel class of tyrosine kinase inhibitors that
exhibit selectivity towards particular receptor tyrosine kinases.
J . Med. Chem. 1998, 41 (14), 2588-2603.
(6) Laird, A. D.; Vajkoczy, P.; Shawver, L. K.; Thurnher, A.; Liang,
C.; Mohammadi, M.; Schlessinger, J .; Ullrich, A.; Hubbard, S.
R.; Blake, R. A.; Fong, A. T.; Strawn, L. M.; Sun, L.; Tang, C.;
Hawtin, R.; Tang, F.; Shenoy, N.; Hirth, K. P.; McMahon, G.;
Cherrington, J . M. SU6668 is a potent antiangiogenic and
antitumor agent that induces regression of established tumors.
Cancer Res. 2000, 60, 4152-4160.
2
-fluorophenyl)[6-methoxy-7-(1-methylpiperidin-4-yl-
1
0
methoxy)quinazolin-4-yl]amine or ZD6474) also im-
proved the solubility of 12g. However, this substitution
increased cytotoxicity and decreased metabolic stability
because of N-demethylation (data not shown). The much
less basic morpholino analogue 12h is also much less
soluble at pH 6, though it has good solubility at pH 2.
The basic heteroaromatic analogues 12i and 12j have
generally poor solubility and sometimes reduced cellular
potency (12j in Table 1).
(
7) Sun, L.; Tran, N.; Liang, C.; Tang, F.; Rice, A.; Schreck, R.;
Waltz, K.; Shawver, L. K.; McMahon, G.; Tang, C. Design,
synthesis, and evaluations of substituted 3-[(3- or 4-carboxy-
ethylpyrrol-2-ylmethylidenyl]indolin-2-ones as inhibitors of VEGF,
FGF, and PDGF receptor tyrosine kinases. J . Med. Chem. 1999,
42, 5120-5130.
(
8) Mendel, D. B.; Laird, A. D.; Xin, X.; Louie, S. G.; Christensen,
J . G.; Li, G.; Schreck, R. E.; Abrams, T. J .; Ngai, T. J .; Lee, L.
B.; Murray, L. J .; Carver, J .; Chan, E.; Moss, K. G.; Haznedar,
J . O.; Sukbuntherng, J .; Blake, R. A.; Sun, L.; Tang, C.; Miller,
T.; Shirazian, S.; McMahon, G.; Cherrington, J . M. In vivo anti-
tumor activity of SU11248, a novel tyrosine kinase inhibitor
targeting VEGF and PDGF receptors: Determination of a
pharmacokinetic/pharmacodynamic relationship. Clin. Cancer
Res. 2003, 9, 327-337.
To assess the protein binding properties for 5b and
1
2b, cellular IC50 values were measured in the presence
or absence of serums. Compared to 5b, the cellular
potency of 12b against PDGF-induced proliferation is
less likely to be affected by the presence of serum
protein (Table 2). In this regard, 12b was still a very
potent inhibitor of PDGF-Râ while 5b became inactive
in the presence of a high level of serum.
(
9) Fong, A. T. T.; Shawver, L. K.; Sun, L.; Tang, C.; App, H.; Powell,
T. J .; Kim, Y. H.; Schreck, R.; Wang, X. Y.; Risau, W.; Ullrich,
A.; Hirth, K. P.; McMahon, G. SU5416 is a potent and selective
inhibitor of the vascular endothelial growth factor receptor (Flk-
1
/KDR) that inhibits tyrosine kinase catalysis, tumor vascular-
In conclusion, 12b possesses the best overall profile
in terms of inhibitory potency against the VEGF-R2 and
ization, and growth of multiple tumor types. Cancer Res. 1999,
59, 99-106.