Development of Next Generation Cell-Permeable Peptide Inhibitors for the Oncological Target MAGE-A4

Melanoma-associated antigen A4 (MAGE-A4) is a cancer/testis antigen (CTA) that interacts with the E3 ubiquitin ligase RAD18 to enhance DNA damage tolerance in tumor cells. Here, we report the structure-guided optimization of a previously reported potent but cell-impermeable cyclic peptide, called MTP-1. Building off our previous peptide inhibitor efforts, we aimed to develop next-generation peptide inhibitors with significantly improved cell permeability. Through systematic structure–activity relationship (SAR) studies employing an mRNA display site-saturation mutagenesis library (SSML) and strategic scaffold optimization with modified cyclization strategy, we developed JWP24, the first cell-permeable peptide inhibitor of MAGE-A4. Evaluation across multiple assays demonstrates intracellular target engagement, maintained binding potency, and exhibits no cytotoxicity at effective concentrations. This study provides a valuable framework for transforming potent but larger, macrocyclic peptide inhibitors into cell-permeable probes. The work presented here demonstrates progress toward further establishing MAGE-A4 as a chemically tractable oncology target.