RT Journal Article
SR Electronic
T1 Proteolysis Targeting Chimeras With Reduced Off-targets
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.11.18.468552
DO 10.1101/2021.11.18.468552
A1 Tuan M. Nguyen
A1 Arghya Deb
A1 Praveen Kokkonda
A1 Vedagopuram Sreekanth
A1 Praveen K. Tiwari
A1 Veronika Shoba
A1 Sophia Lai
A1 Santosh K. Chaudhary
A1 Jaron A. M. Mercer
A1 Max Jan
A1 David R. Liu
A1 Benjamin L. Ebert
A1 Amit Choudhary
YR 2021
UL http://biorxiv.org/content/early/2021/11/20/2021.11.18.468552.abstract
AB Proteolysis Targeting Chimeras (PROTACs), a class of heterobifunctional molecules that recruit target proteins to E3 ligases, have gained traction for targeted protein degradation. However, pomalidomide, a widely used E3 ligase recruiter in PROTACs, can independently degrade other targets, such as zinc-finger (ZF) proteins, that hold key functions in normal development and disease progression. This off-target degradation of pomalidomide-based PROTACs raises concerns about their therapeutic applicability and long-term side effects. Therefore, there is a crucial need to develop rules for PROTAC design that minimize off-target degradation. In this study, we developed a high-throughput platform that interrogates the off-target degradation of ZF domains and discovered, using this platform, that PROTACs with the current design paradigm induce significant degradation of several ZF proteins. To identify new rules for PROTAC design, we generated a rationalized library of pomalidomide analogs with distinct exit vector modifications on the C4 and C5 positions of the phthalimide ring and profiled their propensities for ZF protein degradation. We found that modifications on the C5 position with nucleophilic aromatic substitution (SNAr) reduce off-target ZF degradation. We applied our newfound design principles on a previously developed ALK oncoprotein-targeting PROTAC and generated PROTACs with enhanced potency and minimal off-target degradation. We envision the reported off-target profiling platform and pomalidomide analogs will find utility in design of specific PROTACs.Competing Interest StatementBroad Institute has filed a patent application including the work described herein.