PROTAC design based on first crystal structures of JAK2 kinase domain in complex with ruxolitinib and baricitinib.
PROTACs targeting JAKs are efficacious in vivo in CRLF2r ALL; the most effective degrade multiple targets including JAKs, IKZF1 and GSPT1
CRLF2-rearranged (CRLF2r) acute lymphoblastic leukemia (ALL) comprises over half of Philadelphia chromosome-like (Ph-like) ALL, is associated with poor outcome in children and adults. Overexpression of CRLF2 results in activation of JAK-STAT and parallel signaling pathways in experimental models, but existing small molecule inhibitors of Janus kinases show variable and limited efficacy. Here we evaluated the efficacy of proteolysis-targeting chimeras (PROTACs) directed against Janus kinases. Solving the structure of type I JAK inhibitors ruxolitinib and baricitinib bound to the JAK2 tyrosine kinase domain enabled the rational design and optimization of multiple series of cereblon (CRBN)-directed JAK PROTACs utilizing derivatives of JAK inhibitors, linkers and CRBN-specific molecular glues. The resulting JAK PROTACs were evaluated for target degradation, and activity tested in a panel of leukemia/lymphoma cell lines and xenograft models of kinase-driven ALL. Multiple PROTACs were developed that degraded Janus kinases and potently killed CRLF2-rearranged cell lines, the most active of which also degraded the known CRBN neosubstrate GSPT1, and suppressed proliferation of CRLF2-rearranged ALL in vivo. While dual JAK/GSPT1-degrading PROTACs were most potent, development and evaluation of multiple PROTACs in an extended panel of xenografts identified a potent JAK2-degrading GSPT1-sparing PROTAC that demonstrated efficacy in the majority of the kinase-driven xenografts which were otherwise unresponsive to type I JAK inhibitors. Together, these data show the potential of JAK-directed protein degradation as a therapeutic approach in JAK-STAT-driven ALL, and highlight the interplay of Janus kinase and GSPT1 degradation activity in this context.