Bioorganic & Medicinal Chemistry Letters
The optimization of aminooxadiazoles as orally active inhibitors
of Cdc7
a
a
a
a
Paul E. Harrington a, , Matthew P. Bourbeau , Christopher Fotsch , Michael Frohn , Alexander J. Pickrell ,
Andreas Reichelt a, Kelvin Sham a, Aaron C. Siegmund a, Julie M. Bailis b, Tammy Bush c, Sonia Escobar d,
Dean Hickman e, Scott Heller c, Faye Hsieh e, Jessica N. Orf b, Minqing Rong f, Tisha San Miguel d,
Helming Tan g, Leeanne Zalameda d, John G. Allen a
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a Medicinal Chemistry, Amgen Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA
b Oncology Research, Amgen Inc., 1120 Veterans Blvd., South San Francisco, CA 94080, USA
c Oncology Research, Amgen Inc., 360 Binney Street, Cambridge, MA 02142, USA
d Discovery Technologies, Amgen Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA
e Pharmacokinetics and Drug Metabolism, Amgen Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA
f Oncology Research, Amgen Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA
g Pharmaceutics, Amgen Inc., One Amgen Center Drive, Thousand Oaks, CA 91320, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of aminooxadiazoles was optimized for inhibition of Cdc7. Early lead isoquinoline 1 suffered from
Received 21 August 2013
Revised 16 September 2013
Accepted 18 September 2013
Available online 25 September 2013
modest cell potency (cellular IC50 = 0.71 lM measuring pMCM2), low selectivity against structurally
related kinases, and high IV clearance in rats (CL = 18 L/h/kg). Extensive optimization resulted in azain-
dole 26 (Cdc7 IC50 = 1.1 nM, pMCM2 IC50 = 32 nM) that demonstrated robust lowering of pMCM2 in a
mouse pharmacodynamic (PD) model when dosed orally. Modifications to improve the pharmacokinetic
profile of this series were guided by trapping experiments with glutathione in rat hepatocytes.
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
Cdc7
MCM2
Kinase inhibitor
Azaindole
Cancer
Cell division cycle 7 (Cdc7) is a serine/threonine protein kinase
that plays a pivotal role in the initiation of DNA replication.1 Dur-
ing the S phase of DNA replication, Cdc7 is activated by binding to
the regulatory subunit Dbf4.2 The Cdc7/Dbf4 complex phosphory-
lates one or more minichromosome maintenance complex (MCM2-
7) proteins leading to unwinding of double stranded DNA. The
essential role that Cdc7 plays in S phase entry and DNA replication
has been studied extensively in tumor cells. Cdc7 is overexpressed
in some human tumor cell lines, such as breast, lung, and colon
cancers.3–5 Small interfering RNA (siRNA) knockdown of Cdc7
results in p53 independent apoptotic cell death in tumor cell lines.6
In the same paper,6 siRNA knockdown of Cdc7 in normal cells was
shown to arrest growth reversibly and not cause cell death, sug-
gesting that normal tissue might be spared during cancer treat-
ment with a Cdc7 inhibitor. Furthermore, others have shown that
small molecule Cdc7 inhibitors slow the growth of human tumor
cell lines in mouse xenograft models.7–9 These results spurred
interest in the drug discovery community to develop a small mol-
ecule inhibitor of Cdc7 for use as a single agent or in combination
with chemotherapy.10–17
We recently disclosed a series of N-substituted azaindoles18 and
trisubstituted thiazoles19 as potent inhibitors of Cdc7. Isoquinoline
1 (Fig. 1) emerged as an early lead from a parallel effort to discover
new chemotypes. The chemical matter leading to 1 was identified
through high throughput screening of analogs from our protein
kinase B (PKB) program.20 Compounds were evaluated in an enzy-
matic assay that measured inhibition of Cdc7/Dbf4 biochemical
activity and a cellular assay in HCT-116 cells that measured phos-
phorylation of MCM2 at serine 53.21 Although 1 displayed good
potency in the Cdc7 assay21 (IC50 = 17 nM), it had several deficiencies
that were the focus of SAR efforts. First, potency in the cellular
pMCM2 assay21 (IC50 = 0.71
lM) was modest. Second, the rat IV
pharmacokinetics (PK) of 1 was very poor with an exceptionally
high clearance (CL = 18 L/h/kg) and a very short mean residence
time (MRT) of 0.2 h. We chose to first examine replacements for
the isoquinoline, without changing the aminooxadiazole core and
benzyl amine, with the goal of improving potency and PK. The
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Corresponding author. Tel.: +1 805 313 5564; fax: +1 805 480 1337.
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.