Title page for ETD etd-05152011-184133


Type of Document Master's Thesis
Author Lu, Hao
Author's Email Address haolu@vt.edu
URN etd-05152011-184133
Title Understanding Non-viral Nucleic Acid Delivery Vehicles with Different Charge Centers and Degradation Profiles
Degree Master of Science
Department Chemistry
Advisory Committee
Advisor Name Title
Theresa M. Reineke Committee Chair
Crawford, Daniel T. Committee Member
Santos, Webster L. Committee Member
Keywords
  • Guanidine
  • Biodegradable Polymer
  • Non-viral Nucleic Acid Delivery
  • Self-degradable
Date of Defense 2011-05-10
Availability unrestricted
Abstract
Different structures of non-viral cationic polymer delivery vehicles, including charge center type, molecular weight and degradability, could significantly affect toxicity, release of nucleic acid and transfection efficiency.

Poly(glycoamidoamine)s (PGAAs) contained different carbohydrate and secondary amine moieties and showed high transfection efficiency to different cell lines in a nontoxic manner. The “proton sponge hypothesis” has attempted to relate the buffering capacity to endosomal release of polyethylenimine (PEI) based polyplexes, which could contribute to high transfection efficiency. Secondary amine structures rendered PGAAs buffering capacity around physiological pH. To test the feasibility of the mechanism for PGAAs, new no buffering capacity guanidine or methylguanidine containing poly(glycoamidoguanidine)s (PGAGs) were synthesized. PGAGs formed stable polyplexes with pDNA from N/P (# secondary amine or guanidine group on polymer backbone / # phosphate group on pDNA backbone) ratio 3. PGAG based polyplexes expressed low cytotoxicity and were internalized by 90% of cells at N/P 25. Furthermore, two PGAG based polyplexes showed higher transfection efficiency from N/P 5 to 30 than their PGAA based analogs. These data suggested the low transfection could be due to the difficulties to release pDNA from polyplexes; also, the “proton sponge theory” could not explain the higher transfection efficiency by some PGAGs.

Degradation of delivery vehicles could potentially release pDNA in cells and increase transfection efficiency. PGAAs degraded rapidly at physiological conditions and the proposed mechanism was amide hydrolysis. Typically, amide groups are stable and hydrolyze slowly in absence of enzyme. Different models mimicking PGAAs were synthesized to study the fast hydrolysis. Amide groups showed asymmetric hydrolysis. Different hydrolysis behaviors suggested neighboring group participation of two terminal groups to induce rapid amide hydrolysis. These new models could potentially be used to design new polymer delivery vehicles with various degradation profiles.

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