Development and Mechanistic Evaluation of Cathepsin B‐Activatable Peptide‐Drug Conjugate PROTACs for Targeted Protein Degradation in Malignant Tumors: Advancing Precision Protein Degradation Therapeutics in Oncology

Abstract Since the advent of proteolysis targeting chimeras (PROTACs), both small molecule PROTACs and peptide‐based PROTACs have exhibited distinct advantages. However, PROTACs still exist issues such as poor membrane permeability, low bioavailability, and potential off‐target effects. The development of novel PROTACs represents an innovative approach to addressing these challenges. Here, a class of novel PDC‐PROTACs is designed and modified with cell‐penetrating peptides and tumor‐targeting peptides, which incorporate a Cathepsin B‐sensitive sequence to enhance the targeting of PROTACs to tumor cells and ensure their controlled release within these cells. Subsequent verification employing HPLC‐MRM‐MS technology confirms that the utilization of functional peptides and enzyme‐sensitive linkers significantly augments the intracellular concentration of PDC‐PROTACs, demonstrating improved transmembrane delivery efficiency. Compared to HIF‐S or HIF‐IMA , the enzyme‐responsive PDC‐PROTACs Cyclo‐A7R‐RRR‐GFLG‐HIF‐S and Cyclo‐C9C‐R‐GFLG‐HIF‐IMA demonstrate enhanced anti‐proliferation activity, target protein degradation, and pro‐apoptotic effects. In vivo studies indicate that Cyclo‐A7R‐RRR‐GFLG‐HIF‐S exhibits favorable therapeutic efficacy and safety profile in the U87MG xenograft mouse model. In summary, this research culminates in the creation of novel enzyme‐responsive PDC‐PROTACs, which leverage the elevated expression of specific enzymes in tumor cells to promote their release, thus enabling the degradation of target proteins. The innovative enzyme‐responsive PDC‐PROTACs hold considerable promise for tumor therapy.