2-Hydroxy-4,6-diamino-[1,3,5]triazines: A novel class of VEGF-R2 (KDR) tyrosine kinase inhibitors

Article abstract of DOI:10.1021/jm049372z

2-Hydroxy-4,6-diamino-[1,3,5]triazines are described which are a novel class of potent inhibitors of the VEGF-R2 (flk-1/KDR) tyrosine kinase. 4-(Benzothiazol-6-ylamino)-6-(benzyl-isopropyl-amino)-[1,3,5]triazin-2-ol (14d) exhibited low nanomolar potency i

Full text of DOI:10.1021/jm049372z

J. Med. Chem. 2005, 48, 1717-1720
1717
Scheme 1a
2-Hydroxy-4,6-diamino-[1,3,5]triazines: A
Novel Class of VEGF-R2 (KDR) Tyrosine
Kinase Inhibitors
Nand Baindur, Naresh Chadha, Benjamin M. Brandt,
Davoud Asgari, Raymond J. Patch,
Celine Schalk-HiHi, Theodore E. Carver,
Ioanna P. Petrounia, Christian A. Baumann,
Heidi Ott, Carl Manthey, Barry A. Springer, and
Mark R. Player*
Drug Discovery, Johnson & Johnson Pharmaceutical
Research and Development, L.L.C., 8 Clarke Drive,
Cranbury, New Jersey 08512
Received August 4, 2004
Abstract: 2-Hydroxy-4,6-diamino-[1,3,5]triazines are de-
scribed which are a novel class of potent inhibitors of the
VEGF-R2 (flk-1/KDR) tyrosine kinase. 4-(Benzothiazol-6-
ylamino)-6-(benzyl-isopropyl-amino)-[1,3,5]triazin-2-ol (14d)
exhibited low nanomolar potency in the in vitro enzyme
inhibition assay (IC50 ) 18 nM) and submicromolar inhibitory
activity in a KDR-induced MAP kinase autophosphorylation
assay in HUVEC cells (IC50 ) 280 nM), and also demonstrated
good in vitro selectivity against a panel of growth factor
receptor tyrosine kinases. Further, 14d showed antiangiogenic
activity in an aortic ring explant assay by blocking endothelial
outgrowths in rat aortas with an IC50 of 1 µM.
a
Conditions: (i) 6-aminobenzothiazole, DIEA, THF, 30 min; (ii)
N-methylbenzylamine, DIEA, THF; (iii) 95% TFA in DCM, 60 °C,
4 h; (iv) cumylamine, DIEA, THF; (v) hydroxylamine‚HCl, DIEA,
EtOH, 75 °C, 16 h.
properties in vivo. In this letter, we describe a novel
scaffold, composed of a 2-hydroxy-4,6-diamino-[1,3,5]-
triazine nucleus.
The syntheses of certain 2-hydroxy-4,6-diamino-[1,3,5]-
triazines has been recently described by two different
research groups. In a study of 2,4,6-triamino-[1,3,5]-
triazine inhibitors of the Erm family of methyltrans-
ferases (Erm), the 2-hydroxy-4,6-diamino-[1,3,5]triazine
analogue was synthesized and found to be inactive
(Ki > 100 uM for Erm-AM inhibition), in contrast to the
corresponding 2-amino analogue (Ki ) 8 µM for Erm-
AM inhibition).21 In a different study of a series of 2,4,6-
triamino-[1,3,5]triazines as inhibitors of cholesterol
ester transfer protein (CETP), the corresponding 2-hy-
droxy-4,6-diamino-[1,3,5]triazine analogue was pre-
pared.22 The triazine analogue showed only 11% inhi-
bition of CETP at 50 µM, while the corresponding
hydrazone exhibited a moderate degree of potency (IC50
) 10 µM for CETP inhibition).
Previous approaches for the synthesis of 2-hydroxy-
4,6-diamino-[1,3,5]triazines have utilized solution-phase
methodologies that involve consecutive nucleophilic
displacement of cyanuric chloride. Acid-induced hy-
drolysis (6 N aqueous HCl reflux)21 or base-induced
hydrolysis (aqueous NaOH reflux)22 of the final chlorine
then affords the hydroxytriazine in low to moderate
yields. The present triazines appear to exist in a mixture
of enol and amide forms.23 We found that solution-phase
synthesis of this series (Scheme 1) proceeds most readily
by initial treatment of cyanuric chloride with the
moderately nucleophilic aromatic amine, 6-aminoben-
zothiazole, which gives access to the dichloro-arylamino-
triazine (1) followed by treatment with the more nu-
cleophilic secondary amine, N-methylbenzylamine.
Finally, hydrolysis of the last chlorine was accomplished
by heating with aqueous TFA to yield the desired
hydroxytriazine analogue (3) in relatively good yield.
Treatment of 1 with cumylamine afforded the 2-chloro
analogue (4), which could be further converted to the
Angiogenesis is the process by which new capillaries
form from the established vasculature, and it is neces-
sary for tumors to grow beyond a certain size.1 The
growth of new vessels supplies oxygen, nutrients, and
growth factors to small tumors and removes the waste
products of metabolism. There is also an association
between tumor vascularization and metastatic capacity
of a given tumor.2 Thus, new means to retard angio-
genesis have shown promise as potential cancer thera-
pies. The angiogenic phenotype is also featured in a
variety of pathological states, including diabetic retin-
opathy,3 osteoarthritis,4 and rheumatoid arthritis.5
The vascular endothelial growth factor (VEGF, per-
meability factor) is a primary regulator of physiological
new vessel formation, particularly in wound healing,6
embryogenesis,7 and the female reproductive system.8
It has also been shown to play a direct role in pathologi-
cal neovascularization.9 VEGF recognition, by the high-
affinity receptor tyrosine kinases VEGF-R2 (flk-1/KDR)
and VEGF-R1 (flt-1), is a key mitogenic stimulus
resulting in endothelial cell proliferation, migration, and
vascular permeability.10 KDR is expressed selectively
in endothelial cells.
Disruption of VEGF signaling secondary to capture
of VEGF11 or its receptor12 with antibody has been
demonstrated to retard angiogenesis and inhibit tumor
growth. A number of small molecule structural classes,13
e.g. indolin-2-ones,14 phthalazines,15 quinolinones,16 imi-
dazopyridines,17 benzimidazoles,18 pyridines,19 and
quinazolines,20 have been disclosed as potent inhibitors
of KDR in vitro or have demonstrated antiangiogenic
* To whom correspondence should be addressed. Phone: (609) 655-
6950. Fax: (609) 655-6930. E-mail: mplayer@prdus.jnj.com.
10.1021/jm049372z CCC: $30.25 © 2005 American Chemical Society
Published on Web 11/09/2004

1718 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 6
Letters
Table 1. In Vitro and Cellular Activities (µM) of Hydroxytriazines against KDR
a Data represent the mean (3 determinations) of the concentration needed to inhibit enzyme activity by 50% (IC50 value; µM) at 10 µM
ATP. b Data represent the mean from three independent experiments of the concentration needed to inhibit the growth factor induced
phosphorylation of p42/44 MAP kinase by 50% (IC50 value; µM) by the methods detailed in the Supporting Information.
Scheme 2a
corresponding hydroxylamine (5) by treatment with
hydroxylamine hydrochloride. The 2-methoxy analogue
(8) could be accessed by an alternative synthetic route
starting from the commercially available 4,6-dichloro-
2-methoxy-[1,3,5]triazine (6). Thus, treatment of 6 with
6-aminobenzothiazole forms 1-(6-amino-benzothiazolyl)-
3-chloro-5-methoxytriazine (7) and when followed by
cumylamine yields 8 in relatively good yield.
In order to rapidly develop an SAR around 3 that
would explore the diversity of the two amine substitu-
tions on the triazine ring, an alternative synthetic
a
strategy was desired which would provide facile access
to a wide range of analogues (Scheme 2). Wang resin
was treated with an excess of cyanuric chloride in
anhydrous THF in the presence of DIEA to obtain the
resin-bound dichlorotriazine (9); this in turn underwent
nucleophilic displacement of a chlorine with aromatic
amines (10a-c) in anhydrous THF at room temperature
to yield the resin-bound monochlorotriazines (11a-c).
Reaction utilizing a second set of amines (12a-d) under
more forcing conditions (heating at 90 °C in anhydrous
dioxane for 16 h) led to displacement of the final chlorine
to yield the resin-bound diamino-triazines (13a-f).
Finally, the desired hydroxytriazines (14a-f) were
obtained by cleavage using TFA/DCM conditions (50%
TFA/DCM, 3 h).
Conditions: (i) cyanuric chloride, DIEA, THF; (ii) arylamines
RNHR′ (10a-c), DIEA, THF, rt, 18 h; (iii) amines R′′NHR′′′ (12a-
d), dioxane, 90 °C, 16 h; (iv) 50% TFA in DCM, rt, 3 h.
4- and 6-positions using aryl- and alkylamines at both
positions. The compounds were initially evaluated for
their inhibitory activity toward KDR using a fluores-
cence polarization competitive immunoassay, and the
IC50 values were computed (Table 1). At the outset, the
structural requirements at the 2-position were exam-
ined. A H-bond donor at the 2-position such as hydroxy
(14a) was found to be a key feature. Methylation of the
2-hydroxyl (8) or replacement with Cl (4) resulted in a
greater than 50-fold loss of inhibitory potency. However,
a 2-position hydroxylamine substitution (5) retained
activity both in vitro and in the cell-based assay. At the
4-position, arylamines were found to be virtually es-
sential for inhibitory activity.24 Among the various
Over 1000 2-hydroxy-4,6-diaminotriazines were pre-
pared using this solid-phase approach, varying both the

Letters
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 6 1719
Table 3. Effect on Vessel Outgrowth in Rat Aortic Ring Assaya
Table 2. IC50s (µM) for the Inhibition of the Kinase Activities
of KDR and Related Tyrosine Kinasesa by Hydroxytriazines (5,
14a,c,d)
compd
concn, µM
total tube length ( SEM
control
14d
n/a
30
3
2060 ( 369
36.3 ( 12.9
348 ( 124
1503 ( 227
kinase
5
14a
14c
14d
KDR
Tie-2
FGFR-1
c-fms
c-src
â-IRK
PDGF-R2
0.040
0.7
0.035
1.0
0.049
0.40
>12.5
0.018
0.180
14d
14d
0.3
1.8
>12.5
>12.5
a The total length (arbitrary units) of capillary-like sprouts
emanating outward from individual aortic rings was measured
following 7 days of culture in collagen gels containing 5 ng/mL
VEGF. Values are the mean and SEM from six aortic rings.
14.0
>12.5
4.3
3.1
4.0
2.5
>12.5
>12.5
1.8
4.3
0.42
3.9
0.55
11 ( 5
8.0
>12.5
a IC50 values are the average of 3 determinations. In all cases,
the standard error was found to be less than 2% of the average
value, with the exception of the inhibition of IRK by compound
14d as shown.
arylamine substitutions, those displaying the best ac-
tivities were 6-aminobenzothiazole (14a) and 6-amino-
indazole (14f). Of these, 14a (IC50 ) 35 nM) was
superior to 14f (IC50 ) 240 nM) in vitro. In addition,
14f showed substantially weaker activity in the cell-
based assay (IC50 >20 µM), possibly as a result of
impaired cellular permeability. N-Methylation of 6-ami-
nobenzothiazole (14e) reduced activity almost 40-fold
(IC50 ) 1.6 µM) in the in vitro assay. Interestingly,
elaboration of the N-substituent to methyl (3) and then
to ethyl (14c) each resulted in approximately 7-fold
enhancement of the inhibitory potency, possibly as a
reflection of favorable hydrophobic binding. An N-
isopropyl substituent (14d) was subsequently found to
be optimal and afforded the most potent compound in
this series with an in vitro IC50 of 18 nM as well as
potency in the cell-based HUVEC assay (IC50 ) 289 nM).
In the absence of a tertiary amine at the 6-position,
rotationally hindered benzylamines such as cumylamine
(14a) afforded good potency.
In order to assess the selectivity of these KDR
inhibitors, inhibitory potencies were measured for com-
pounds 5, 14a, 14c, and 14d against a panel of tyrosine
kinases. C-fms and PDGF-R2 are members of the class
III family of receptor tyrosine kinases, which includes
KDR. Tie-2 is another receptor tyrosine kinase that is
involved in vascular development and is potentially an
important target for blocking angiogenesis at a second-
ary stage.25 For the compounds tested, selectivity for
KDR relative to the kinases in Table 2 was always at
least 9-fold or greater. Compound 14d in particular was
found to have a potentially desirable selectivity profile,
yielding significant inhibition against Tie-2 kinase only.
This could be advantageous, as a dual KDR/Tie-2 kinase
inhibitor may exhibit enhanced antiangiogenic activity
in vivo.
Figure 1. Inhibition of vessel formation in a rat aortic ring
model (14d). A light micrograph (original 10×) of representa-
tive control and 14d-treated aortic rings. Emanating from
control rings were capillary-like sprouts, which are absent in
the 14d-treated cultures.
kinase phosphorylation in DMSO-treated control cells.
Several of the best compounds inhibited VEGF-induced
MAP kinase activation with IC50s in the submicromolar
range (Table 1). Because these compounds had some
modest activity toward other kinases in vitro, we
proceeded to evaluate cell-based specificity of the com-
pounds toward a diverse set of receptor tyrosine kinases.
We tested compound activity in cells expressing the
human insulin receptor (CHO-IR), mouse FGFR1 (NIH
3T3 fibroblasts), and the human EGF receptor (A431).
The compounds were evaluated for inhibition of IR,
FGF-R1, and EGFR induced phosphorylation of MAP
kinase in cells. This was done using a phosphorylation
specific ELISA method that directly detects growth
factor induced phosphorylation of MAP kinase in cells
grown, treated, and fixed to 96-well tissue culture dishes
as described in the Supporting Information.26 Using this
quantitative method compounds 5, 12a, 12c, and 12d
all displayed IC50s of greater than 10 µM in the cell-
based counterscreens for EGF-R, IR, and FGF-R1
activation. Although compounds 5, 12a, 12c, and 12d
displayed some activity toward the insulin receptor
kinase in enzyme studies, the compounds lack signifi-
cant cell-based insulin receptor inhibition. This is most
likely due to either the cell permeability of the com-
pounds or, because the compounds are ATP competitive,
the increased ATP concentration in the cellular environ-
ment compared to the in vitro enzyme assay condition
(1-2 mM and 10 µM respectively). Additionally, these
results suggest that compounds 5, 12a, 12c, and 12d
are not significantly affecting key kinases in the MAP
kinase pathway downstream of most receptor tyrosine
kinases.
Since an inhibitor of KDR must bind to an intra-
cellular kinase domain, the compounds were evaluated
in a cellular assay using a HUVEC cell line, which
endogenously expresses KDR. Inhibition of KDR activity
in HUVECs was monitored through analysis of KDR-
induced MAP kinase phosphorylation. Low passage (3-
12) HUVEC cells were grown to confluence, serum
starved, treated with compound, or DMSO and stimu-
lated with VEGF. HUVECs were then lysed, and the
lysates were subjected to SDS-PAGE followed by
quantitative immunoblotting with a phospho-specific
MAP kinase (pERK1/2) antibody. Inhibition of KDR-
induced MAP kinase phosphosphorylation was quanti-
fied as a percent of the total VEGF-stimulated MAP
An initial evaluation of the antiangiogenic activity of
this novel class of KDR inhibitors was carried out using
an aortic ring explant assay.27 In this assay, rings cut
from aortas were embedded and cultured in collagen
gels and gave rise over 7 days to endothelial outgrowths
resembling microcapillary sprouts. Nicosia et al. re-
ported previously that at least 70% of the angiogenic

1720 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 6
Letters
(14) 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-carboxyeth-
ylpyrrol-2-yl)methylidenyl]indolin-2-ones as inhibitors of VEGF,
FGF, and PDGF receptor tyrosine kinases. J. Med. Chem. 1999,
42, 5120-5130. Sun, L.; Tran, N.; Liang, C,; Hubbard, S.; Tang,
F.; Lipson, K.; Schreck, R.; Zhou, Y.; McMahon, G.; Tang, C.
Identification of substituted 3-[(4,5,6,7-tetrahydro-1H-indol-2-
yl)methylene]-1,3-dihydroindol-2-ones as growth factor receptor
inhibitors for VEGF-R2 (Flk-1/KDR), FGF-R1, and PDGF-Râ
tyrosine kinases. J. Med. Chem. 2000, 43, 2655-2663.
(15) Bold, G.; Altmann, K. H.; Frei, J.; Lang, M.; Manley, P. W.;
Traxler, P.; Wietfeld, B.; Bruggen, J.; Buchdunger, E.; Cozens,
R.; Ferrari, S.; Furet, P.; Hofmann, F.; Martiny-Baron, G.;
Mestan, J.; Rosel, J.; Sills, M.; Stover, D.; Acemoglu, F.; Boss,
E.; Emmenegger, R.; Lasser, L.; Masso, E.; Roth, R.; Schlachter,
C.; Vetterli, W. New anilinophthalazines as potent and orally
well absorbed inhibitors of the VEGF receptor tyrosine kinases
useful as antagonists of tumor-driven angiogenesis. J. Med.
Chem. 2000, 43, 2310-2323.
response can be blocked by neutralizing antibody against
VEGF,27 underscoring the importance of VEGF signal-
ing in this assay system. We found that compound 14d
blocked vascular sprout formation by 83% and 27% at
3 and 0.3 µM, respectively (Table 3 and Figure 1). The
approximate IC50 for compound 14d in the aortic ring
explant assay (1 µM) is somewhat higher than the IC50
for blocking HUVEC proliferation (0.28 µM). This may
reflect potential species differences in compound potency
or compound binding to collagen gels.
In summary, we have described a novel series of
2-hydroxy-4,6-diamino-[1,3,5]triazines, which are potent
inhibitors of KDR kinase, and which have comparable
selectivities to other reported small molecule inhibitors
in preclinical development.
(16) Fraley, M. E.; Arrington, K. L.; Buser, C. A.; Ciecko, P. A.; Coll,
K. E.; Fernandes, C.; Hartman, G. D.; Hoffman, W. F.; Lynch,
J. J.; McFall, R. C.; Rickert, K.; Singh, R.; Smith, S.; Thomas,
K. A.; Wong, B. K. Optimization of the indolyl quinolinone class
of KDR kinase inhibitors: effects of 5-amido- and 5-sulfonamido-
indolyl groups on pharmacokinetics and hERG binding. Bioorg.
Med. Chem. Lett. 2004, 14, 351-355.
Supporting Information Available: Experimental pro-
cedures and spectra for the research described in this letter.
This material is available free of charge via the Internet at
http://pubs.acs.org.
(17) Wu, Z.; Fraley, M. E.; Bilodeau, M. T.; Kaufman, M. L.; Tasber,
E. S.; Balitza, A. E.; Hartman, G. D.; Coll, K. E.; Rickert, K.;
Shipman, J.; Shi, B.; Sepp-Lorenzino, L.; Thomas, K. A. Design
and synthesis of 3,7-diarylimidazopyridines as inhibitors of the
VEGF-receptor KDR. Bioorg. Med. Chem. Lett. 2004, 14, 909-
912.
(18) Bilodeau, M. T.; Cunningham, A. M.; Koester, T. J.; Ciecko, P.
A.; Coll, K. E.; Huckle, W. R.; Hungate, R. W.; Kendall, R. L.;
McFall, R. C.; Mao, X.; Rutledge, R. Z.; Thomas, K. A. The
discovery of N-(1,3-thiazol-2-yl)pyridin-2-amines as potent in-
hibitors of KDR kinase. Bioorg. Med. Chem. Lett. 2003, 13,
2485-2488.
(19) Bilodeau, M. T.; Rodman, L. D.; McGaughey, G. B.; Coll, K. E.;
Koester, T. J.; Hoffman, W. F.; Hungate, R. W.; Kendall, R. L.;
McFall, R. C.; Rickert, K. W.; Rutledge, R. Z.; Thomas, K. A.
The discovery of N-(1,3-thiazol-2-yl)pyridin-2-amines as potent
inhibitors of KDR kinase. Bioorg. Med. Chem. Lett. 2004, 14,
2941-2945.
(20) Hennequin, L. F.; Thomas, A. P.; Johnstone, C.; Stokes, E. S.;
Pl_, P. A.; Lohmann, J. J.; Ogilvie, D. J.; Dukes, M.; Wedge, S.
R.; Curwen, J. O.; Kendrew, J.; Lambert-van der Brempt, C.
Design and structure-activity relationship of a new class of
potent VEGF receptor tyrosine kinase inhibitors. J. Med. Chem.
1999, 42, 5369-5389. Hennequin, L. F.; Stokes, E. S.; Thomas,
A. P.; Johnstone, C.; Ple´, P. A.; Ogilvie, D. J.; Dukes, M.; Wedge,
S. R.; Kendrew, J.; Curwen, J. O. Novel 4-anilinoquinazolines
with C-7 basic side chains: design and structure activity
relationship of a series of potent, orally active, VEGF receptor
tyrosine kinase inhibitors. J. Med. Chem. 2002, 45, 1300-1312.
(21) Hadjuk, P. J.; Dinges, J.; Schkeryantz, J. M.; Janowick, D.;
Kaminski, M.; Tufano, M.; Augeri, D. J.; Petros, A.; Nienaber,
V.; Zhong, P.; Hammond, R.; Coen, M.; Beutel, B.; Katz, L.;
Fesik, S. W. Novel Inhibitors of Erm Methyltransferases from
NMR and Parallel Synthesis. J. Med. Chem. 1999, 42, 3852-
3859.
References
(1) Carmeliet, P.; Jain, R. K. Angiogenesis in cancer and other
diseases. Nature 2000, 401, 249-257. Ferrara, N. VEGF and
the quest for tumour angiogenesis factors. Nat. Rev. Cancer
2002, 2, 795-803. Manley, P. W.; Martiny-Baron, G.; Schlaeppi,
J. M.; Wood, J. M. Therapies directed at vascular endothelial
growth factor. Expert Opin. Invest. Drugs 2002, 11, 1715-1736.
(2) McDonnell, C. O.; Hill, D. A.; McNamara, D. A.; Walsh, T. N.;
Bouchler-Hayes, D. J. Tumor micrometastases: the influence
of angiogenesis. Eur. J. Surg. Oncol. 2000, 26, 105-115.
(3) Patel, N.; Sun, L.; Moshinsky, D.; Chen, H.; Leahy, K. M.; Le,
P.; Moss, K. G.; Wang, X.; Rice, A.; Tam, D.; Laird, A. D.; Yu,
X.; Zhang, Q.; Tang, C.; McMahon, G.; Howlett, A. A selective
and oral small molecule inhibitor of vascular epithelial growth
factor receptor (VEGFR)-2 and (VEGFR)-1 inhibits neovascu-
larization and vascular permeability. J. Pharmacol. Exp. Ther.
2003, 306, 838-845.
(4) Enomoto, H.; Inoki, I.; Komiya, K.; Shiomi, T.; Ikeda, E.; Obata,
K.; Matsumoto, H.; Toyama, Y.; Okada, Y. Vascular endothelial
growth factor isoforms and their receptors are expressed in
human osteoarthritic cartilage. Am. J. Pathol. 2003, 162, 171.
(5) Clavel, G.; Bessis, N.; Boissier, M. C. Recent data on the role of
angiogenesis in rheumatoid arthritis. Joint Bone Spine 2003,
70, 321-326.
(6) Tonnesen, M. G.; Feng, X.; Clark, R. A. Angiogenesis in wound
healing. J. Invest. Dermatol. Symp. Proc. 2000, 5, 40-46.
(7) Zhou, Y.; Genbacev, O.; Fisher, S. J. The human placenta
remodels the uterus by using a combination of molecules that
govern vasculogenesis or leukocyte extravasation. Ann. N.Y.
Acad. Sci. 2003, 995, 73-83.
(8) Reynolds, L. P.; Grazul-Bilska, A. T.; Redmer, D. A. Angiogenesis
in the female reproductive organs: pathological implications. Int.
J. Exp. Pathol. 2002, 83, 151-163.
(9) Kim, K. J.; Li, B.; Winer, J.; Armanini, M.; Gillett, N.; Phillips,
H. S.; Ferrara, N. Inhibition of vascular endothelial growth
factor-induced angiogenesis suppresses tumour growth in vivo.
Nature 1993, 362, 841-844.
(10) Ferrara, N.; Gerber, H. P. The role of vascular endothelial growth
factor in angiogenesis. Acta Haematol. 2001, 106, 148-156.
(11) Kabbinavar, F.; Hurwitz, H. I.; Fehrenbacher, L.; Meropol, N.
J.; Novotny, W. F.; Lieberman, G.; Griffing, S.; Bergsland, E.
Phase II, randomized trial comparing bevacizumab plus fluo-
rouracil (FU)/leucovorin (LV) with FU/LV alone in patients with
metastatic colorectal cancer. J. Clin. Oncol. 2003, 21, 60-65.
Yang, J. C.; Haworth, L.; Sherry, R. M.; Hwu, P.; Schwartzen-
truber, D. J.; Topalian, S. L.; Steinberg, S. M.; Chen, H. X.;
Rosenberg, S. A. A randomized trial of bevacizumab, an anti-
vascular endothelial growth factor antibody, for metastatic renal
cancer. New Engl. J. Med. 2003, 349, 427-434.
(12) Prewett, M.; Huber, J.; Li, Y.; Santiago, A.; O’Connor, W.; King,
K.; Overholser, J.; Hooper, A.; Pytowski, B.; Witte, L.; Bohlen,
P.; Hicklin, D. J. Antivascular endothelial growth factor receptor
(fetal liver kinase 1) monoclonal antibody inhibits tumor angio-
genesis and growth of several mouse and human tumors. Cancer
Res. 1999, 59, 5209-5218.
(13) For a review of KDR inhibitors, see: Boyer, S. J. Small molecule
inhibitors of KDR (VEGFR-2) kinase: an overview of structure
activity relationships. Curr. Top. Med. Chem. 2002, 2, 973-1000.
(22) Xia, Y.; Mirzai, B.; Chackalamannil, S.; Czarniecki, M.; Wang,
S.; Clemmons, A.; Ahn, H.-S.; Boykow, G. C. Substituted 1,3,5-
Triazines as Cholesterol Ester Transfer Protein Inhibitors.
Bioorg. Med. Chem. Lett. 1996, 6, 919-922.
(23) See Supporting Information (IR spectra 3) and the following:
Socrates, G. Infrared Characteristic Group Frequencies; George
Wiley and Sons: New York, 1994; pp 136-137.
(24) Data not shown.
(25) Thurston, G. Role of Angiopoietins and Tie receptor tyrosine
kinases in angiogenesis and lymphangiogenesis. Cell Tissue Res.
2003, 314, 61-8.
(26) Versteeg, H. H.; Nijhuis, E.; van der Brink, G. R.; Evertzen, M.;
Pynaert, G. N.; van Deventer, S. J. H.; Coffer, P. J.; Peppelen-
bosch, M. P. A new phosphospecific cell-based ELISA for the p42/
44 mitogen-activated (MAPK), p38 MAPK and protein kinase
B and cAMP-response-element binding protein. Biochem. J.
2000, 350, 717-722.
(27) Nicosia, R. F.; Lin, Y. J.; Hazelton, D.; Qian, X. Endogenous
regulation of angiogenesis in the rat aorta model. Role of
vascular endothelial growth factor. Am. J. Pathol. 1997, 151,
1379-1386.
JM049372Z