Proposed Allosteric Inhibitors Bind to the ATP Site of CK2α

CK2α
is a ubiquitous, well-studied kinase that is a target
for small-molecule inhibition, for treatment of cancers. While many
different classes of adenosine 5′-triphosphate (ATP)-competitive
inhibitors have been described for CK2α, they tend to suffer
from significant off-target activity and new approaches are needed.
A series of inhibitors of CK2α has recently been described as
allosteric, acting at a previously unidentified binding site. Given
the similarity of these inhibitors to known ATP-competitive inhibitors,
we have investigated them further. In our thorough structural and
biophysical analyses, we have found no evidence that these inhibitors
bind to the proposed allosteric site. Rather, we report crystal structures,
competitive isothermal titration calorimetry (ITC) and NMR, hydrogen–deuterium
exchange (HDX) mass spectrometry, and chemoinformatic analyses that
all point to these compounds binding in the ATP pocket. Comparisons
of our results and experimental approach with the data presented in
the original report suggest that the primary reason for the disparity
is nonspecific inhibition by aggregation.