Entry into a new class of protein kinase inhibitors by pseudo ring design

Article abstract of DOI:10.1016/j.bmcl.2007.12.041

A pyrimidin-4-yl-urea motif forming a pseudo ring by intramolecular hydrogen bonding has been designed to mimic the pyrido[2,3-d]pyrimidin-7-one core structure of a well-established class of protein kinase inhibitors. Potent inhibition of a number of prot

Full text of DOI:10.1016/j.bmcl.2007.12.041

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 18 (2008) 897–900
Entry into a new class of protein kinase inhibitors
by pseudo ring design
Pascal Furet,* Giorgio Caravatti, Vito Guagnano, Marc Lang,
Thomas Meyer and Joseph Schoepfer
Novartis Institutes for BioMedical Research, WKL-136.P.12, CH-4002 Basel, Switzerland
Received 23 November 2007; revised 14 December 2007; accepted 19 December 2007
Available online 14 January 2008
Abstract—A pyrimidin-4-yl-urea motif forming a pseudo ring by intramolecular hydrogen bonding has been designed to mimic the
pyrido[2,3-d]pyrimidin-7-one core structure of a well-established class of protein kinase inhibitors. Potent inhibition of a number of
protein kinases was obtained with the first prototype compound synthesized to probe the design concept.
Ó 2007 Elsevier Ltd. All rights reserved.
Designing molecular mimics of established lead com-
pounds is one of the major approaches in medicinal
chemistry to generate new biologically active chemo-
types. In such efforts, one aims at introducing the chem-
ical functions conferring the desired biological activity
in new, different, molecular frameworks. One thus ex-
pects to obtain molecules reaching the level of activity
of the reference lead compound and presenting some
additional benefit such as easier synthetic access, better
patentability or different physico-chemical properties
that might turn out advantageous in the drug develop-
ment phase.
protein kinase inhibitors.4,5 In particular, attachment of
an aryl moiety in position 6 of the ring system provides
broadly active tyrosine kinase inhibitors exemplified by
PD 166285.6 Thus, our interest in identifying new tyro-
sine kinase inhibitor scaffolds led us to take PD 166285
(Fig. 1) as a template for the design of prototype mimics.
Previous efforts in this direction are reported in the liter-
ature.7 They consist of replacing one of the rings of the
core pyrido[2,3-d]pyrimidin-7-one system by a similar
Cl
6
Pseudo six-membered rings resulting from the formation
of an intramolecular hydrogen bond in a planar conju-
gated moiety of a molecule are considered to be partic-
ularly stable. They are often observed in crystal
structures of organic molecules.1 Replacing real rings
by such pseudo rings is a new and non-conventional
strategy to try to mimic established lead compounds.
Recently, we have reported the discovery of new classes
of potent VEGF-R and EGF-R tyrosine kinase inhibi-
tors following this approach.2,3 In this letter, we report
another example of successful pseudo ring design ap-
plied to the search for new kinase inhibitor scaffolds.
O
N
N
Cl
O
N
N
H
N
PD166285
Cl
O
H
N
N
N
N
Cl
O
N
H
N
H
Molecules based on a pyrido[2,3-d]pyrimidin-7-one core
structure constitute one of the most prominent classes of
1
Keywords: Kinase; Scaffold morphing; Pseudo ring.
*
Figure 1. Chemical structures of PD 166285 and prototype compound
1.
Corresponding author. Tel.: +41 61 696 79 90; fax: +41 61 696 48
70; e-mail: pascal.furet@novartis.com
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.12.041

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P. Furet et al. / Bioorg. Med. Chem. Lett. 18 (2008) 897–900
H-bond
R3
R3
H
5
N
N
N
N
R1
R1
O
N
N
H
O
N
N
H
N
1
R2
R2
Figure 2. Pseudo ring design concept.
tural Database using a pyrimidin-4-yl-urea substructure
query returned seven molecules with available crystal
structure coordinates.8 Encouragingly, we noticed that
in all of them the pyrimidin-4-yl-urea motif adopts the
pseudo cyclic conformation presenting an intramolecu-
lar hydrogen bond between the pyrimidine ring and
the urea group. We then performed ab initio calcula-
tions on the model compound shown in Figure 3. The
difference in energy between the extended and pseudo
cyclic conformations of this model compound was com-
puted in the gas phase and using a water solvation mod-
el. In both cases, the pseudo cyclic conformation was
calculated to be more stable, strengthening our motiva-
tion to synthesize the envisaged prototype compound.9
H
N
H
N
O
H
N
H
N
N
N
O
N
N
A
B
Figure 3. Model compound studied by ab initio calculations. Using the
DFT-B3LYP/6-31G** method, the pseudo cyclic conformation B is
predicted to be more stable than the extended conformation A.
heterocyclic moiety, in a traditional manner. We decided
to follow a more risky but novel approach to achieve
this objective. As illustrated in Figure 2, one of our de-
sign ideas was to move the nitrogen atom in position 1
of the pyrimidine ring of PD 166285 to position 5 and
replacing the pyrimidone ring by a urea moiety able to
form an intramolecular hydrogen bond with this nitro-
gen atom. The pseudo six-membered ring thus obtained
looked to us as an excellent mimic of the pyrimidone
ring and the synthesis of compound 1 as a first proto-
type for the designed pyrimidinyl urea scaffold was
envisaged.
The pyrimidinyl urea 1 was prepared in two steps from
commercially available starting materials (Scheme 1). 6-
Chloro-pyrimidin-4-ylamine and 4-(2-diethylamino-eth-
oxy)-phenylamine were condensed in n-butanol at
120 °C using 1 equiv of hydrochloric acid in dioxane.
The intermediate 2 was further condensed with 1,3-di-
chloro-2-isocyanato-benzene in 1,3-dimethyl-3,4,5,6-tet-
rahydro-2(1H)-pyrimidinone
temperature.
(DMPU)
at
room
Compound 1 was tested in biochemical assays measur-
ing its ability to inhibit the catalytic activity of various
recombinant protein kinases.10 The experimental data
are reported in Table 1. To our satisfaction, several tyro-
sine kinases such as c-Src, EGF-R, c-Abl, and Tie-2
were inhibited in the submicromolar range giving sup-
port to our design concept. Based on the known binding
mode of pyrido[2,3-d]pyrimidin-7-one kinase inhibi-
Before engaging in the synthesis of 1, we sought some
experimental and computational evidence that the pseu-
do cyclic conformation required to mimic the pyr-
ido[2,3-d]pyrimidin-7-one bicyclic system was indeed
stable and would not prohibit binding to a kinase active
site because of a too high energy cost to pay for confor-
mational adaptation. A search in the Cambridge Struc-
H2N
H
N
Cl
N
N
i
N
O
+
N
N
N
O
NH2
NH2
2
Cl
N
H
O
N
N
Cl
N
N
O
Cl
H
N
NH
ii
O
1
Cl
Scheme 1. Synthesis of compound 1. Reagents and conditions: (i) 4 N HCl in dioxane, n-butanol, 120 °C; (ii) solvent: N,N0-dimethyl-N,N0-propylene
urea, rt, yield: 37%.

P. Furet et al. / Bioorg. Med. Chem. Lett. 18 (2008) 897–900
899
Table 1. IC50 values (lM) of compound 1 in enzymatic assays and
nature of gate keeper residue
possessing a bulky gate keeper residue is inhibited by 1
at a concentration as high as 10 lM. This is the case
for the FLT3, JAK2, and c-Met kinases, for instance,
which present a phenylalanine, a methionine, and a leu-
cine, respectively, at the gate keeper position. In con-
trast, all the kinases tested having a threonine or a
residue of similar size (valine or isoleucine) at this posi-
tion are inhibited by 1 in the micromolar or submicrom-
olar range.
a
Kinase
IC50
Gate keeper
c-Src
EGF-R
c-Abl
0.066
0.38
0.25
0.57
0.93
0.96
0.30
1.4
Thr
Thr
Thr
Val
Thr
Val
Ile
FGFR-1
c-Kit
KDR
Tie-2
PDGFR-b
B-raf V599E
EphB4
P38
Thr
Thr
Thr
Thr
Phe
Leu
Met
Met
Phe
Met
Met
Leu
In conclusion, the results presented here give an addi-
tional illustration that employing a pseudo ring resulting
from the formation of an intramolecular hydrogen bond
is a valuable novel approach for mimicking a real ring in
a lead compound. Prototype 1 has given us an entry in a
new class of protein kinase inhibitors of potential inter-
est for a variety of therapeutic targets.14–16
0.15
0.43
0.35
>10
>10
>10
>10
>10
>10
>10
>10
FLT3
c-Met
IGF1-R
JAK2
CDK2
PKA
PKB
Axl
Acknowledgment
a All IC50 values represent averages of at least three experimental
determinations.
We thank K. Ucci-Stoll for her technical assistance.
References and notes
1. Bilton, C.; Allen, F. H.; Shields, G. P.; Howard, J. A. K.
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W. D.; Steinkampf, R. W.; Driscoll, D. L.; Nelson, J. M.;
Elliott, W. L.; Roberts, B. J.; Stoner, C. L.; Vincent, P.
W.; Dykes, D. J.; Panek, R. L.; Lu, G. H.; Major, T. C.;
Dahring, T. K.; Hallak, H.; Bradford, L. A.; Showalter,
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5. VanderWel, S. N.; Harvey, P. J.; McNamara, D. J.;
Repine, J. T.; Keller, P. R.; Quin, J., 3rd; Booth, R. J.;
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L. J. Med. Chem. 2005, 48, 2371.
6. Panek, R. L.; Lu, G. H.; Klutchko, S. R.; Batley, B. L.;
Dahring, T. K.; Hamby, J. M.; Hallak, H.; Doherty, A.
M.; Keiser, J. A. J. Pharmacol. Exp. Ther. 1997, 283, 1433.
7. For recent examples, see (a) Cao, J.; Fine, R.; Gritzen, C.;
Hood, J.; Kang, X.; Klebansky, B.; Lohse, D.; Mak, C.
C.; McPherson, A.; Noronha, G.; Palanki, M. S. S.;
Pathak, V. P.; Renick, J.; Soll, R.; Zeng, B.; Zhu, H.
Bioorg. Med. Chem. Lett. 2007, 17, 5812; (b) McDermott,
L. A.; Simcox, M.; Higgins, B.; Nevins, T.; Kolinsky, K.;
Smith, M.; Yang, H.; Li, J. K.; Chen, Y.; Ke, J.; Mallalieu,
N.; Egan, T.; Kolis, S.; Railkar, A.; Gerber, L.; Luk, K.-
C. Bioorg. Med. Chem. Lett. 2005, 13, 4835; (c) Sabat, M.;
VanRens, J. C.; Brugel, T. A.; Maier, J.; Laufersweiler, M.
J.; Golebiowski, A.; De, B.; Easwaran, V.; Hsieh, L. C.;
Rosegen, J.; Berberich, S.; Suchanek, E.; Janusz, M. J.
Bioorg. Med. Chem. Lett. 2006, 16, 4257.
8. These structures have the following entry codes in the
Cambridge Structural Database: BPCTHA, CNBPCT,
KPRCGM20, PCMTRE10, PCTRIB10, PUCGLR10,
and PURTRE.
9. The ab initio calculations were performed in the program
Jaguar (Jaguar, version 6.0, Schro¨dinger, LLC, New
York, NY, 2005) using the DFT-B3LYP method with
Figure 4. Model of compound 1 (green) docked in the ATP pocket of
the c-Abl kinase (yellow). A CPK representation is given to the gate
keeper residue, a threonine for this kinase. The canonical hydrogen
bonds to the hinge segment are indicated by dashed lines.
tors11 a model of 1 docked in the ATP pocket of one of
these kinases (c-Abl) was constructed (Fig. 4).12 In the
model, the dichlorophenyl moiety of our pyrimidin-4-
yl-urea mimic makes favorable van der Waals contacts
with the hydrophobic back pocket of the ATP binding
site, in particular with the threonine gate keeper resi-
due.13 The model suggests a very tight fit between the
inhibitor and the enzyme in this region of the ATP pock-
et. As a consequence, an adverse steric clash between the
compound and the gate keeper residue is expected if this
is significantly larger than a threonine. In full agreement
with this notion, none of the kinases of the Table 1 panel

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P. Furet et al. / Bioorg. Med. Chem. Lett. 18 (2008) 897–900
the 6-31G** basis set and doing full geometry optimiza-
Hendrickson, T.; Still, W. C. J. Comput. Chem. 1990, 11,
tion. A preliminary systematic conformational analysis of
the model compound by molecular mechanics varying all
torsion angles identified the extended and pseudo cyclic
conformations A and B as the low energy conformations
of this system. In the subsequent quantum mechanical
calculations, B was computed to be more stable than A: by
3.2 kcal/mol in the gas phase and by 0.5 kcal/mol applying
the water solvation model available in Jaguar.
440.
13. The gate keeper is the main residue determining the
selectivity of ATP site directed kinase inhibitors. Named
like this because it controls the access to the hydrophobic
back pocket of the ATP binding site, it is located at the
beginning of the hinge segment, the amino acid stretch
that connects the N- and C-terminal domains of a kinase.
In c-Abl, this residue is Thr315. For recent papers
discussing the role of the gate keeper residue in determin-
ing kinase inhibitor selectivity, see for instance (a) Furet,
P.; Bold, G.; Meyer, T.; Roesel, J.; Guagnano, V. J. Med.
Chem. 2006, 49, 4451; (b) Vieth, M.; Higgs, R. E.;
Robertson, D. H.; Shapiro, M.; Gragg, E. A.; Hemmerle,
H. Biochim. Biophys. Acta., Proteins Proteomics 2004,
1697, 243.
14. Following the publication of our patent application
covering this new class of kinase inhibitors,15 scientists
at Procter and Gamble Pharmaceuticals have reported
their independent efforts leading to the discovery of
related urea compounds as potent inhibitors of the Lck
kinase.16
15. Ding, Q.; Gary, N. S.; Li, B.; Liu, Y.; Sim, T.; Uno, T.;
Zhang, G.; Pissot Soldermann, C.; Breitenstein, W.; Bold,
G.; Caravatti, G.; Furet, P.; Guagnano, V.; Lang, M.;
Manley, P. W.; Schoepfer, J.; Spanka, C. WO 2006/
000420. Chem. Abstr. 2006, 144, 108347.
16. Maier, J. A.; Brugel, T. A.; Sabat, M.; Golebiowski, A.;
Laufersweiler, M. J.; VanRens, J. C.; Hopkins, C. R.; De,
B.; Hsieh, L. C.; Brown, K. K.; Easwaran, V.; Janusz, M.
J. Bioorg. Med. Chem. Lett. 2006, 16, 3646.
10. The protein kinase inhibition assays used in this study are
described in the following references (a) Garcia-Echever-
ria, C.; Pearson, M. A.; Marti, A.; Meyer, T.; Mestan, J.;
Zimmermann, J.; Gao, J.; Brueggen, J.; Capraro, H.-G.;
Cozens, R.; Evans, D. B.; Fabbro, D.; Furet, P.; Porta, D.
G.; Liebetanz, J.; Martiny-Baron, G.; Ruetz, S.; Hof-
mann, F. Cancer Cell 2004, 5, 231; (b) Furet, P.; Bold, G.;
Meyer, T.; Roesel, J.; Guagnano, V. J. Med. Chem. 2006,
49, 4451.
11. Nagar, B.; Bornmann, W. G.; Pellicena, P.; Schindler, T.;
Veach, D. R.; Miller, W. T.; Clarkson, B.; Kuriyan, J.
Cancer Res. 2002, 62, 4236.
12. Compound 1 was docked in the crystal structure of the
kinase domain of c-Abl in complex with a pyrido[2,3-
d]pyrimidin-7-one inhibitor (PD 173955) published in
reference 11 (PDB entry code 1M52). Compound 1 was
aligned on PD 173955 in the ATP pocket according to our
mimetism concept. Then, PD 173955 was removed and the
resulting complex energy minimized in Macromodel using
the AMBER/H2O/GBSA force field. Macromodel
Mohamadi, F.; Richards, N. G. J.; Guida, W. C.;
Liskamp, R.; Lipton, M.; Caufield, C.; Chang, G.;