Compound B1(Hangzhou Medical College)

Proteolysis targeting chimeras (PROTACs) offer a promising therapeutic approach by leveraging the ubiquitin–proteasome system (UPS) to degrade target proteins. These heterobifunctional molecules consist of a target protein ligand, an E3 ligase ligand, and a linker. Among the limited E3 ligase ligands available, cereblon (CRBN) ligands are the most widely used. However, the stability and racemization of current chiral CRBN ligands pose challenges for developing CRBN‐recruiting PROTAC therapeutics. Herein, a partial PROTAC library is reported incorporating an aldehyde motif and various linkers into previously developed achiral phenyl dihydrouracil CRBN ligands, which offer improved stability and eliminate racemization issues. This library enables the rapid generation of fully functional PROTACs targeting Bruton's tyrosine kinase (BTK) by coupling the aldehyde motif with a hydrazide‐containing BTK ligand. Initial HiBiT assay (Promega) screening identifies nine hits capable of significant BTK degradation, with compound B1 emerging as the most potent degrader. A stable amide bioisostere, AM‐B1, is further developed, which induces significant antiproliferation and BTK degradation. Mechanistic studies confirm BTK degradation via the UPS. This study highlights the development of an achiral CRBN‐based partial PROTAC library and demonstrates a two‐stage strategy for rapid PROTAC development against BTK.

Hematopoietic progenitor kinase 1 (HPK1, MAP4K1) is a promising target for immune-oncology therapy. It has been recently demonstrated that loss of HPK1 kinase activity can enhance T cell receptor (TCR) signaling. However, many essential functions mediated by the HPK1 scaffolding role are still beyond the reach of any kinase inhibitor. Proteolysis targeting chimera (PROTAC) has emerged as a promising strategy for pathogenic proteins degradation with the characteristics of rapid, reversible, and low-cost versus RNA interference or DNA knock-out technology. Herein we first disclosed the design, synthesis, and evaluation of a series of thalidomide-based PROTAC molecules and identified B1 as a highly efficient HPK1 degrader with DC₅₀ value of 1.8 nM. Further mechanism investigation demonstrated that compound B1 inhibits phosphorylation of the SLP76 protein with IC₅₀ value of 496.1 nM, and confirmed that B1 is a bona fide HPK1-PROTAC degrader. Thus, this study provides a basis for HPK1 degraders development and the candidate could be used as a potential chemical tool for further investigation of the kinase-independent signaling of HPK1 in TCR.