Design, Synthesis, and Biological Evaluation of Lysine-Stapled Peptide Inhibitors of p53-MDM2/MDMX Interactions with Potent Antitumor Activity In Vivo

We introduce novel lysine-stapled peptide inhibitors targeting p53-MDM2/MDMX interactions. Leveraging the model peptides PDI (LTFEHYWAQLTS) and PMI-M3 (LTFLEYWAQLMQ) as starting points, a series of lysine-stapled analogues were designed and synthesized. Through in vitro cell assay screening, two lead compounds, SPDI-48-T₁ and SPMI-48-T₃, were identified for their excellent antiproliferation activity. Fluorescence polarization assays revealed that both compounds exhibited strong binding affinities against MDM2 and MDMX, ascertained by K_d values within the low micromolar spectrum. Further characterization of SPDI-48-T₁ and SPMI-48-T₃ demonstrated that SPDI-48-T₁ possessed superior cell permeability and serum stability. Notably, SPDI-48-T₁ displayed a dose-dependent suppression of tumor growth in an HCT116 xenograft mouse model. Our findings indicate that SPDI-48-T₁ holds promise as a lead compound for further development as an Anticancer agent by modulating p53-MDM2/MDMX interactions. Additionally, this study also proved that the lysine stapling strategy may serve as a robust approach for generating peptide ligands targeting Other protein-protein interactions.