cb500355c_si_002.mpg (1.47 MB)
Discovery of Inter-Domain StabilizersA Novel Assay System for Allosteric Akt Inhibitors
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posted on 2015-01-16, 00:00 authored by Zhizhou Fang, Jeffrey R. Simard, Dennis Plenker, Hoang D. Nguyen, Trang Phan, Patrik Wolle, Stefan Baumeister, Daniel RauhIn
addition to the catalytically active kinase domain, most kinases
feature regulatory domains that govern their activity. Modulating
and interfering with these interdomain interactions presents a major
opportunity for understanding biological systems and developing novel
therapeutics. Therefore, small molecule inhibitors that target these
interactions through an allosteric mode of action have high intrinsic
selectivity, as these interactions are often unique to a single kinase
or kinase family. Here we report the development of iFLiK (interface-Fluorescent
Labels in Kinases), a fluorescence-based assay that can monitor such
interdomain interactions. Using iFLiK, we have demonstrated selective
detection of allosteric Akt inhibitors that induce an inactive closed
conformation unique to Akt. This methodology easily distinguished
small molecule allosteric inhibitors from classic ATP-competitive
inhibitors. Screening an in-house compound library with iFLiK, we
were able to identify novel compounds with a scaffold that has not
been previously described for allosteric Akt inhibitors.