1096
M. J. Mulvihill et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1091–1097
Table 7. PK in mice for compounds 6a.4, 6f.1, and 6i.1
suitable in vivo PK and PD effects in accordance with
tumor growth inhibition.
Compound
6a.4
6f.1
6i.1
Cmax at 50 mg/kg, po (lM)
AUC0ꢀ1 at 50 mg/kg, po (ng h/mL)
Oral bioavailability (%F)
t1/2 (h) at 1 mg/kg iv
0.86
1756
47
2.22
1323
34
2.85
3926
47
Acknowledgments
1.30
0.63
2.29
We thank Professor Victor Snieckus for consultation; Ms.
Viorica M. Lazarescu and Dr. Minghui Wang for analyt-
ical support; Dr. Arno Steinig, Mrs. Yingchuan Sun, and
Dr. Jonathan Williams for their synthetic efforts on the 1-
three animals were sacrificed at designated timepoints,
and blood samples were collected in EDTA following
cardiac puncture. Following centrifugation at 1500g
for 10 min, plasma samples were extracted by protein
precipitation with methanol and analyzed by HPLC-
MS/MS (PE Sciex API 3000 triple quadrupole mass
spectrometer, Applied Biosystems). Drug levels were
quantitated against spiked calibration samples pre-
pared in control mouse plasma. Pharmacokinetic
parameters were calculated by non-compartmental
analysis using the median (n = 3) concentration at each
timepoint. This analysis showed the compounds to be
orally bioavailable with the major parameters present-
ed in Table 7.
(3-benzyloxyphenyl)-imidazo[1,5-a]pyrazines
series
which were not included in this manuscript; and Dr.’s Arl-
indo Castelhano, Neil Gibson, and Lee Arnold for their
managerial support and guidance.
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In order to determine selectivity of this class of com-
pounds, representative compound 6i.1 was profiled for
inhibitory activity against 15 additional purified protein
kinases from the tyrosine and serine/threonine kinase
families. Protein kinase assays were performed at Up-
state Inc. (Charlottesville, VA) by a radiometric method
(KinaseProfiler service). Inhibition of protein kinases by
compound 6i.1 was assessed at 10 lM using recombi-
nant purified enzymes in the presence of ATP concentra-
tion approximating the respective Km value for each
form of the enzyme. Less than 50% inhibition at
10 lM of the compound was noted for the following en-
zymes: Ab1, Cdk2/CyclinA, Cdk2/CyclinE, Chk2, CK2,
c-Raf, Fes, IKK-b, MAPK2, p70S6K, PDGFR-b,
PDK1, PKA, PKBa, and PKCa. Inhibition of IR was
similar to that observed for IGF-IR, reinforcing the
high homology between the ATP binding domains of
these proteins.
In summary, a series of novel 8-amino-1,3-disubstitut-
ed-imidazo[1,5-a]pyrazines was designed and synthe-
sized as IGF-IR inhibitors.
A
combination of
systematic medicinal chemistry exploration and model-
ing-based design-guided modifications of lead com-
pound 6a.4 around the benzyloxy tether, the 8-amino
group, and the cycloalkyl moiety at C3 of the imidazo-
pyrazine ring ultimately led to a new series of potent
inhibitors, the most interesting of which are the amin-
omethylcyclohexyl and cyclobutyl analogs (compounds
6i and 6l, respectively). Emphasis on addressing the po-
tential metabolic liabilities associated with the benzyl-
oxyphenyl moiety has led our efforts to a search for
alternate C1 tethers. In subsequent manuscripts, we
will describe the selection of a more constrained quin-
oline tether, designed through structure-based efforts
derived from IGF-IR and IR X-ray co-crystal struc-
tures with select imidazopyrazines. In addition, we will
discuss efforts to establish selectivity against the highly
homologous insulin receptor as well as demonstrate