3660
T. C. Coombs et al. / Bioorg. Med. Chem. Lett. 23 (2013) 3654–3661
Figure 6. Docking pose for compound 17 in Clk4 homology model: (a) Important hydrogen bond interactions with the hinge region of the enzyme; (b) surface representation
showing hydrophobic pocket. (Graphics prepared using PYMOL.)
Table 3
further evaluated and support the idea that this compound is a
selective Clk/Dyrk inhibitor with adequate solubility, stability,
IC50 values (nM) against Clk and Dyrk kinases for small-molecule inhibitors
and cell permeability to allow it to be used in cell-based biological
studies. Compound 35 (ML315), therefore, represents a comple-
mentary addition to the very small collection of existing Clk/Dyrk
inhibitors (Table 3). Its biochemical profile, when compared to
other inhibitors, should make it a useful biochemical tool, particu-
larly if used in parallel with other inhibitors to dissect Clk/Dyrk
biology.
Compound
Clk1
Clk2
Clk3
Clk4
Dyrk1A
Dyrk1B
35 (ML315)
139
68
59
1522
19
20
231
1902
1648
95
>10,000
6936
>10,000
3000
530
4500
68
39
282
62
>10,000
12
55
40
1156
697
243
136
30
4420
130
TG00339
KH-CB1917
Leucettine L41
NRa
NRa
NRa
NRa
NRa
NRa
18
15
a
NR = not reported.
Acknowledgments
of SYBYL. In this arrangement, compound 17 forms several hydro-
gen bonds with amino acid residues Leu244, Lys191, and Asp325 in
the hinge region of the enzyme. The importance of the hydrogen
bond between the oxygen of the benzodioxole and Leu244 may
be seen when it is compared to analogs in which the dioxole ring
is replaced by either a p-methoxy (10/11) or m-methoxy (12/13)
substituent. In the m-methoxy analogs, the oxygen atom is not in
proximity to Leu244, while in the p-methoxy cases, the methyl
group would be expected to rotate out of the plane of the ortho
C–H groups, positioning the oxygen lone pairs away from the
Leu244 residue and disrupting hydrogen bond formation. These
observations are reflected in the steep drops in inhibitory activity
for compounds 10–13. The same effect is observed for veratrole-
substituted analog 21 for similar reasons. Ring-expanded benzodi-
oxane analog 20, on the other hand, is still a potent Clk4 inhibitor
as the two methylene carbons are held rigidly in the ring, placing
the lone pairs on the para oxygen atom at angles conducive to
hydrogen bond formation with Leu244. Comparing compounds
having the guanidyl core to compounds having the amidinyl core,
the importance of the guanidyl nitrogen as a hydrogen bond accep-
tor with Lys191 may also be seen. In the amidinyl series, a hydro-
gen bond with Lys191 is absent and the compound is significantly
less potent. The surface representation in Figure 6b suggests that
the aromatic ring of the benzylamine moiety sits in a hydrophobic
pocket where van der Waals interactions between the halogenated
benzylamine and the pocket are important for binding.
In conclusion, a new series of aryl-substituted aminopyrimi-
dines with activity against the Clk and Dyrk families of kinases
has been described. Four substitution patterns around the central
pyrimidine were explored, and a number of new compounds were
discovered with activities <100 nM against combinations of Clk1,
Clk2, Clk4, Dyrk1A and Dyrk1B. The most potent agents have activ-
ities <10 nM. Three compounds with different substitution pat-
terns were subjected to DiscoverRXÒ KINOMEscanÒ analysis,
revealing different levels of selectivity within the kinome between
the chemotypes. The off-target pharmacology and in vitro pharma-
cokinetic properties of the most selective of these agents, 35, were
This work was supported by National Institutes of Health
(grants U54 HG005031, RO3 MH084827-01, and U54 MH084681-
01) and the Division of Preclinical Innovation, National Center for
Advancing Translational Sciences of the National Institutes of
Health. The dendrograms depicting kinase binding selectivity were
We thank Benjamin Neuenswander and Patrick Porubsky for com-
pound purification and curation. We thank Arianna Mangravita-
Novo and Layton H. Smith of the Sanford-Burnham Medical
Research Institute at Lake Nona for solubility and PK profiling
measurements.
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