Solid-phase guanidyl-formation triggered peptide stapling and cyclization: A robust platform for improving multiple properties of bioactive peptides

Stapled or cyclic peptides offered advantageous membrane permeability and proteolytic stability to linear counterparts. We herein synthesized a series of S-alkyl-isothiourea building blocks, which could be conveniently coupled with one or two amine groups in both i,i+4 and i,i+7 space located within the peptides, enabling the silver-promoted solid-phase guanidyl-formation (SSG) peptide stapling. The guanidyl stapled peptides, characterized with highly hydrophilic and cationic properties, could be readily labelled by biotin, polyethylene glycol and fluorescent motif using the pre-made functionalized bis-S-alkyl-isothiourea. SSG reactions could be rendered for generating double-stapled peptide, and notably, tri-guanidyl bicyclic peptide with the aid of tri-S-alkyl-isothiourea donor. When compared with hydrocarbon stapled and/or linear peptide, SSG stapled peptides exhibited enhanced proteolytic tolerance, remarkable hydrophilicity, membrane penetration and thus higher in vitro biological activities. Our developed SSG peptide stapling and cyclization strategy served as a meaningful tool in modifying other peptide fragments and offering the potent peptide therapeutic candidates.