Aberrant activation of S6 kinase1 (S6K1) is found in many diseases,including diabetes, aging, and cancer. We developed ATP competitiveorganometallic kinase inhibitors, EM5 and FL772, which are inspiredby the structure of the pan-kinase inhibitor staurosporine, to specificallyinhibit S6K1 using a strategy previously used to target other kinases.Biochemical data demonstrate that EM5 and FL772 inhibit the kinasewith IC50value in the low nanomolar range at 100 μMATP and that the more potent FL772 compound has a greater than 100-foldspecificity over S6K2. The crystal structures of S6K1 bound to staurosporine,EM5, and FL772 reveal that the EM5 and FL772 inhibitors bind in theATP binding pocket and make S6K1-specific contacts, resulting in changesto the p-loop, αC helix, and αD helix when compared tothe staurosporine-bound structure. Cellular data reveal that FL772is able to inhibit S6K phosphorylation in yeast cells. Together, thesestudies demonstrate that potent, selective, and cell permeable S6K1inhibitors can be prepared and provide a scaffold for future developmentof S6K inhibitors with possible therapeutic applications. [ABSTRACT FROM AUTHOR]