title: Sequence segregation improves non-covalent protein delivery.
abstract: The impermeability of the plasma membrane towards large, hydrophilic biomolecules is a major obstacle in their use and development against intracellular targets. To overcome such limitations, protein transduction domains (PTDs) have been used as protein carriers, however they often require covalent fusion to the protein for efficient delivery. In an effort to develop more efficient and versatile biological vehicles, a series of PTD-inspired polyoxanorbornene-based synthetic mimics with identical chemical compositions but different hydrophobic/hydrophilic segregation were used to investigate the role of sequence segregation on protein binding and uptake into Jurkat T cells and HEK293Ts. This series was composed of a strongly segregated block copolymer, an intermediately segregated gradient copolymer, and a non-segregated homopolymer. Among the series, the block copolymer maximized both protein binding and translocation efficiencies, closely followed by the gradient copolymer, resulting in two protein transporter molecules more efficacious than currently commercially available agents. These two polymers were also used to deliver the biologically active Cre recombinase into a loxP-reporter T cell line. Since exogenous Cre must reach the nucleus and retain its activity to induce gene recombination, this in vitro experiment better exemplifies the broad applicability of this synthetic system. This study shows that increasing segregation between hydrophobic and cationic moieties in these polymeric mimics improves non-covalent protein delivery, providing crucial design parameters for the creation of more potent biological delivery agents for research and biomedical applications.
authors.affiliation: Department of Polymer Science & Engineering, University of Massachusetts, Amherst, MA 01003, United States.
authors.forename: Federica
authors.initials: F
authors.lastname: Sgolastra
authors.affiliation: Department of Polymer Science & Engineering, University of Massachusetts, Amherst, MA 01003, United States.
authors.forename: Coralie M
authors.initials: CM
authors.lastname: Backlund
authors.affiliation: Molecular and Cellular Biology Program, University of Massachusetts, Amherst, MA 01003, United States.
authors.forename: E
authors.initials: E
authors.lastname: Ilker Ozay
authors.affiliation: Department of Polymer Science & Engineering, University of Massachusetts, Amherst, MA 01003, United States.
authors.forename: Brittany M
authors.initials: BM
authors.lastname: deRonde
authors.affiliation: Molecular and Cellular Biology Program, University of Massachusetts, Amherst, MA 01003, United States; Department of Veterinary & Animal Sciences, University of Massachusetts, Amherst, MA 01003, United States.
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Minter
authors.affiliation: Department of Polymer Science & Engineering, University of Massachusetts, Amherst, MA 01003, United States; Molecular and Cellular Biology Program, University of Massachusetts, Amherst, MA 01003, United States; Department of Veterinary & Animal Sciences, University of Massachusetts, Amherst, MA 01003, United States. Electronic address: tew@mail.pse.umass.edu.
authors.forename: Gregory N
authors.initials: GN
authors.lastname: Tew
mesh_terms: D001692:Biological Transport; D004337:Drug Carriers / Q000737:chemistry*; D065546:Drug Liberation; D057809:HEK293 Cells; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D019426:Integrases / Q000378:metabolism; D019169:Jurkat Cells; D009636:Norbornanes / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry*; D010539:Permeability; D011108:Polymers / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D000072417:Protein Domains; D011506:Proteins / Q000008:administration & dosage* / Q000737:chemistry* / Q000378:metabolism; D011995:Recombination, Genetic
chemical_list: D004337:Drug Carriers; D009636:Norbornanes; D010455:Peptides; D011108:Polymers; D011506:Proteins; C045073:Cre recombinase; D019426:Integrases
keywords: Cell penetrating peptides; Non-covalent protein delivery; Pep-1; Protein binding; Protein transduction domain mimics; Sequence segregation