Increased Hydrophobic Block Length of PTDMs Promotes Protein Internalization

Presenter: 

Coralie M Backlund

Authors: 

Coralie M. Backlund (1), Federica Sgolastra (1), Ronja Otter (10, Lisa Minter (2,3), Toshihide Takeuchi (4), Shiroh Futaki (4), and Gregory N. Tew (1,2,3)

Author Affiliation: 

1Department of Polymer Science & Engineering, University of Massachusetts, Amherst, MA 01003 2Department of Veterinary & Animal Sciences, University of Massachusetts, Amherst, MA 01003 3Department of Molecular & Cellular Biology, University of Massachusetts, Amherst, MA 01003 4Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan.

Abstract: 

The plasma membrane is a major obstacle in the development and use of biomacromolecules for intracellular delivery applications. Consequently, proteins with intracellular targets represent an enormous, yet under studied avenue for therapeutics. Protein transduction domains (PTDs) have been used to overcome the plasma membrane, but often require covalent conjugation to the protein and can be time consuming to synthesize. Polymers can be used to mimic amino acid moieties in PTDs as they provide a well-controlled platform to vary molecular architecture for structure activity relationship studies. More specifically, ROMP is an ideal synthesis platform as it is extremely functional group tolerant and offers fast, controlled polymerizations with narrow dispersities allowing for facile tunability of the hydrophobic and cationic domains in these PTD mimics. In this study, a series of polyoxanorbornene-based synthetic mimics inspired by PTDs with varying hydrophobic and cationic densities and contents were investigated in vitro to determine their ability to non-covalently transport proteins into multiple cell lines using enhanced green fluorescent protein. The hydrophobic content of each polymer was investigated using an HPLC assessment of their hydrophobic monomers and the overall effect of hydrophobicity on polymer size in solution by DLS. The polymer with the highest hydrophobic content proved to be the most efficient at internalization, while decreasing hydrophobic density and content decreased overall protein delivery efficiency. These results provide crucial design parameters for more effective internalization of biomacromolecules for intracellular delivery applications.