Parallel De Novo Design and Screening of multiple site-specific Cell-Permeable β-Hairpin-based Inhibitors

The primary goal of this project is to integrate advanced techniques for the generation of Permeable β-hairpin-derived Inhibitors (PβhdI) of intracellular targets. Specifically, the approach will combine site recognition and epitope discovery from de novo design with enhanced affinity achieved through phage-display maturation. Additionally, it will improve stability and permeability via interstrand crosslinking (e.g., using bis-electrophiles or disulfides), which will also facilitate phage library diversification. By using these techniques synergistically, the project aims to significantly advance the design of intracellular drugs, potentially broadening the range of available therapeutic options and enhancing the efficacy of targeted treatments. The methodology focuses on the large-scale design and characterization of cell-PβhdI that target multiple binding sites on an intracellular therapeutic target, such as β-catenin. β-Catenin, a key component of the Wnt signaling pathway, is an attractive intracellular target due to its role in various cancers driven by aberrant Wnt signaling. The heterovalent nature of β-catenin, which possesses four distinct binding sites and well-characterized crystal structures, provides an ideal framework for assessing the effectiveness of multiple site-specific binders. Those, combined with the host institution's expertise in studying this oncogene, makes β-catenin an ideal candidate for applying and refining this new methodology.