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Y. Shin et al. / Bioorg. Med. Chem. Lett. 19 (2009) 3344–3347
Table 3
Aryl piperidines
N
N
a
a
Compd
R3
JNK3 IC50
(
lM)
JNK1 IC50
(
lM)
4g
9b
9d
9e
9f
9g
9h
9i
Br
Cl
F
CN
Me
CHF2
Et
0.06
0.08
0.41
0.21
0.53
0.35
1.0
0.09
0.04
0.21
0.23
0.33
0.26
1.4
H
N
Br
Cl
O
O
9j
9k
9l
9m
9n
Propynyl
Bn
OMe
NHAc
NHPh
0.11
4.8
0.62
11.0
>20
0.11
3.2
0.45
5.8
>20
a
Values are means of three experiments. All standard deviation 6 20%.
Given the finding that 3-methyl substitution was slightly more
potency enhancing than 3-chloro (4p vs 4e), future SAR was con-
ducted on the 3-methylsubstituted analogs (Table 3). Additionally,
a spiroketal-piperidine was found to be more potency enhancing
than the terminally substituted piperazine group. While attempts
to replace the bromofuran ring proved challenging, we were able
to replace the bromine atom without complete loss of activity.
The 5-chlorofuran was the most promising substitution with little
affect on potency, however all other substitutions tried, led to a 5–
10-fold drop. This drop in potency did not appear to be a size-re-
lated phenomena given that both smaller and larger groups are
equally less active.
Figure 2. X-ray crystal structure of active site binding 4g (cyan structure) and JNK3
(green). Critical residues and distance between inhibitor amide oxygen atom and
Met 149 amide NH are labeled.
A few compounds from this series were further profiled in a
Acknowledgments
cell-based assay (4f IC50 = 3.3 lM; 4d IC50 = 2.1 lM) and showed
a considerable shift in activity (30–40-fold).24 Whether the shift
arises from the higher ATP concentration in cells or from lack of
cell penetration is still under investigation, but this is not an
uncommon observation.25
We would like to thank Peter Hodder and the Scripps Florida
HTS Group for screening as well as Professor Bill Roush for helpful
discussions. We also thank the beamline staff at SER-CAT (APS 22-
ID) for the data collection help and support in solving the protein
structure. This work was supported by NIH grant U01 NS057153
awarded to P.L.
To help explain the binding mode of inhibitor to enzyme and to
aid in the design of more potent analogs, an X-ray crystal structure
of a compound from this class bound to JNK3 was pursued. Fortu-
nately, 4g provided crystals suitable for X-ray diffraction and the
interactions between ligand and enzyme are shown in Figure 2.26
The inhibitor occupied the ATP binding site, with the bromofuran
ring placed deep within the adenosine binding region. The furan
was up against the gatekeeper Met 146, but interacting only
through van der Waals contacts. The piperazine also formed van
der Waals contacts with Ile 70 and Val 78 and to a lesser extent
it is interacting in a similar fashion with Val 196 and Leu 206.
These are important because they help stabilize the p-loop. The
amide oxygen atom in 4g bound to the backbone hinge (Met
149) in JNK3 and was also close enough to form an electrostatic
interaction with the carboxy main chain oxygen of Glu 147. This
binding mode is not unlike that described for other structurally un-
ique JNK inhibitors in the literature which benefit not from a
strong bidentate interaction with the hinge region of the enzyme,
but from the sum of many weak interactions.27 It may also help
to explain why small perturbations in the molecular structure
greatly affect inhibition.
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In summary, a novel class of piperazine amides were devel-
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a series of potent pan-JNK
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inhibitors inactive against p38 that displayed moderate activity
in a JNK cell-based assay. An X-ray crystal structure with JNK3
revealed an unusual binding mode which may be helpful in
designing more potent analogs. Synthesis and characterization
of these compounds is in progress and will be reported in
due course.