In situ ROPISA synthetic strategy for a β-sheet polypeptide nanoformulation and in vivo biodistribution studies

Polypeptides are protein-like synthetic macromolecules that often require tedious post-polymerization self-assembly to encapsulate drugs or genes. Developing nanotechnology platforms for the on-demand loading of cargoes during synthesis in water would yield directly injectable nanoformulations for immediate healthcare applications. Here, we report one of the first attempts to construct an aqueous-medium-stabilized β-sheet poly(L-tyrosine) block copolymer polypeptide nanoparticle formulation via a ring-opening polymerization-induced self-assembly (ROPISA) process. The synthetic scope of the methodology is further expanded to include the in situ encapsulation of biologically relevant fluorophore molecules, which are purified by a simple dialysis process and directly injected for in vitro and in vivo studies in biological systems. The β-sheet polypeptide nanoformulation is non-toxic to cells at up to 100 μg mL^-1, non-hemolytic to red blood cells, and readily endocytosed across the cell membranes, as substantiated by confocal microscopy imaging. A deep-tissue penetrable near-infrared IR-780 biomarker-loaded nanoformulation is employed for biodistribution studies in live animals (mice). The β-sheet polypeptide formulation is stable under blood circulation, and ex vivo tissue imaging established their accumulation largely in the digestive organs. This approach paves the way for the direct injectable polypeptide nanoformulations in drug delivery.