Exploiting An Asp-Glu "Switch" In Glycogen Synthase Kinase 3 To Design Paralog-Selective Inhibitors For Use In Acute Myeloid Leukemia

Glycogen synthase kinase 3 (GSK3), a key regulatory kinase in the wingless-type MMTV integration site family (WNT) pathway, is a therapeutic target of interest in many diseases. Although dual GSK3 alpha/beta inhibitors have entered clinical trials, none has successfully translated to clinical application. Mechanism-based toxicities, driven in part by the inhibition of both GSK3 paralogs and subsequent beta-catenin stabilization, are a concern in the translation of this target class because mutations and overexpression of beta-catenin are associated with many cancers. Knockdown of GSK3 alpha or GSK3 beta individually does not increase beta-catenin and offers a conceptual resolution to targeting GSK3: paralog-selective inhibition. However, inadequate chemical tools exist. The design of selective adenosine triphosphate (ATP)-competitive inhibitors poses a drug discovery challenge due to the high homology (95% identity and 100% similarity) in this binding domain. Taking advantage of an Asp(133) -> Glu(196) "switch" in their kinase hinge, we present a rational design strategy toward the discovery of paralog-selective GSK3 inhibitors. These GSK3 alpha- and GSK3 beta-selective inhibitors provide insights into GSK3 targeting in acute myeloid leukemia (AML), where GSK3 alpha was identified as a therapeutic target using genetic approaches. The GSK3 alpha-selective compound BRD0705 inhibits kinase function and does not stabilize beta-catenin, mitigating potential neoplastic concerns. BRD0705 induces myeloid differentiation and impairs colony formation in AML cells, with no apparent effect on normal hematopoietic cells. Moreover, BRD0705 impairs leukemia initiation and prolongs survival in AML mouse models. These studies demonstrate feasibility of paralog-selective GSK3 alpha inhibition, offering a promising therapeutic approach in AML.