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Date of Award
Doctor of Philosophy (PhD)
Polymer Science and Engineering
Gregory N. Tew
Biochemistry | Polymer Chemistry
Currently, most of the commercially available therapeutics are all targeting cell surface receptors which constitutes only a small portion of the targets found in the cells. Therefore, reaching intracellular targets would provide many new opportunities to treat various diseases. However, intracellular delivery of therapeutic molecules has always been a challenge due to the poor permeability of cell membrane to large, negatively charged macromolecules and their restricted biodistribution. In the past decades, cell penetrating peptides (CPPs), also known as protein transduction domains (PTDs), are shown to improve the intracellular delivery of bioactive molecules and among the PTDs, arginine-rich peptides are highlighted as the most effective subclass. In the light of this information, using the power of polymer chemistry, protein transduction domain mimics (PTDMs) based on ring opening metathesis polymerization (ROMP) of functionalized oxanorbornene derivatives are aimed to be designed. This thesis demonstrates that these PTDMs can adopt cell penetrating activity and show superior properties compared to peptide analogues (i.e. nonaarginine, R9, Pep-1). The structure-activity relationship is studied by guanidinium functionalized monomers. The impact of number of guanidiniums, density of guanidiniums, molecular length and hydrophobicity on cellular internalization is investigated. Further, the siRNA delivery ability of designed PTDMs is also studied. Efficient downregulation of NOTCH1 protein using PTDM-based non-covalent siRNA delivery system in T cell lines and primary blood cells is demonstrated. Two different structures of PTDMs are studied to understand the structural requirements for an efficient carrier. Apart from in vitro testing of PTDM/siRNA complexes, their size and surface charge are also characterized. Further, PTDM-based siRNA delivery system is used to study the function of NOTCH1 in in vitro in primary human blood cells and as well as in humanized mouse model of graft vs host disease as an in vivo environment. In addition to siRNA delivery, novel protein transporter PTDMs which are inspired by primary amphipathic peptides is introduced. The effects of different functional groups and different block lengths on protein delivery efficiency are studied. Successful delivery of functional proteins is demonstrated using Cre Recombinase and Runx1.d190.
Tezgel, Arife Ozgul, "Protein Transduction Domain Mimics by ROMP and Their Bioactive Cargo Delivery" (2013). Doctoral Dissertations 1896 - February 2014. 452.