Esterase-responsive albumin-binding PROTAC-mediated BRD4 degradation for cancer immunotherapy.

Rationale: Proteolysis-targeting chimeras (PROTACs) represent a powerful therapeutic modality for selective protein degradation but often suffer from poor pharmacokinetics and limited tumor-targeting. To overcome these constraints, we developed albumin-binding BRD4-degrading PROTACs (Alb-TACs) with esterase-cleavable maleimide linkers that hitchhike endogenous albumin and enable esterase-responsive BRD4 degradation in tumors. Methods: Alb-TACs were synthesized by conjugating two esterase-cleavable maleimide linkers, bicyclononyne-polyethylene glycol-maleimide (BCN-PEG Results: Alb-TAC#2 containing the AE-Mal linker exhibited rapid albumin binding, accelerated esterase-responsive activation, and enhanced tumor accumulation compared to ARV-771 and Alb-TAC#1 due to its flexible chemical structure. In the CT26 cell culture system, Alb-TAC#2 efficiently degraded BRD4, resulting in BRD4-deficient cell death. Furthermore, in CT26 tumor-bearing mice, Alb-TAC#2 achieved extensive apoptosis through robust BRD4 degradation, leading to marked downregulation of c-Myc, Bcl-2, and PD-L1. Moreover, Alb-TAC#2 induced hallmarks of ICD (elevated surface CRT, extracellular ATP, and HMGB1) and reprogrammed the tumor microenvironment by enhancing CD8⁺ T cell infiltration, promoting dendritic cell maturation, and reducing regulatory T cell function. Conclusions: This esterase-responsive albumin-binding PROTAC design could overcome pharmacokinetic barriers of conventional BRD4-targeting PROTACs by enhancing tumor-specific delivery and esterase-responsive BRD4 degradation in solid tumors. In summary, esterase-responsive albumin-binding PROTAC is proven as a promising strategy that effectively modulates the pharmacokinetics and therapeutic performance of PROTACs for cancer immunotherapy.