In Vitro Selection of Macrocyclic α/ß3-Peptides against Human EGFR.

Here, we report ribosomal construction of thioether-macrocyclic α/ß3-peptide libraries in which ß-homoglycine, ß-homoalanine, ß-homophenylglycine, and ß-homoglutamine are introduced by genetic code reprogramming. The libraries were applied to the RaPID (Random nonstandard Peptides Integrated Discovery) selection against human EGFR to obtain PPI (protein-protein interaction) inhibitors. The resulting peptides contained up to five ß3-amino acid (ß3AA) residues and exhibited outstanding binding affinity, PPI inhibitory activity, and proteolytic stability, which were attributed to the ß3AAs included in the peptides. This showcase work has demonstrated that the use of such ß3AAs enhances the drug-like properties of peptides, providing a unique platform for the discovery of de novo macrocycles against a protein of interest.

De novo peptide grafting to a self-assembling nanocapsule yields a hepatocyte growth factor receptor agonist.

Lasso-grafting (LG) technology is a method for generating de novo biologics (neobiologics) by genetically implanting macrocyclic peptide pharmacophores, which are selected in vitro against a protein of interest, into loops of arbitrary protein scaffolds. In this study, we have generated a neo-capsid that potently binds the hepatocyte growth factor receptor MET by LG of anti-MET peptide pharmacophores into a circularly permuted variant of Aquifex aeolicus lumazine synthase (AaLS), a self-assembling protein nanocapsule. By virtue of displaying multiple-pharmacophores on its surface, the neo-capsid can induce dimerization (or multimerization) of MET, resulting in phosphorylation and endosomal internalization of the MET-capsid complex. This work demonstrates the potential of the LG technology as a synthetic biology approach for generating capsid-based neobiologics capable of activating signaling receptors.