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Author ORCID Identifier
https://orcid.org/0009-0005-7247-9318
Date Available
4-13-2026
Year of Publication
2026
Document Type
Doctoral Dissertation
Degree Name
Doctor of Philosophy (PhD)
College
Engineering
Department/School/Program
Chemical and Materials Engineering
Faculty
Daniel Pack
Faculty
J. Zach Hilt
Abstract
Gene therapy has promised to revolutionize medicine since the earliest clinical trials, but a lack of delivery methods that are both safe and efficient has hampered progress. Gene therapy with viral vectors is effective at delivering nucleic acids to cells, but is associated with dangerous immunogenic responses and cancer-causing insertions into the genome. Viral vectors are also among the most prohibitively costly therapeutics in medicine. Polymeric vectors avoid these dangers and are made from inexpensive, off-the- shelf materials, but have been hindered by poor efficiency compared to viral vectors.
A benchmark polymer for in vitro nucleic acid delivery has been cationic polyethylenimine (PEI). However, PEI/DNA complexes suffer from instability and poor transfection efficiency in the presence of serum, which has limited clinical applications. A recent development has been the discovery of greatly enhanced transgene expression in the presence of serum by succinylated zwitterionic polyethylenimine/DNA (zPEI/DNA) complexes, but the mechanisms through which zPEI/DNA expression is enhanced are not understood.
This work investigates why zPEI/DNA polyplexes enable robust transgene expression in the presence of serum. Compared to unmodified PEI/DNA polyplexes, zPEI/DNA polyplexes are monodisperse, do not form heterogeneous aggregates in the presence of protein, and exhibit more stable packaging. In HeLa and HEK293 cells, while PEI/DNA polyplexes promiscuously use multiple endocytic mechanisms, are reliant on acidification, and accumulate in the endolysosome, zPEI/DNA polyplexes are selective for lipid raft-mediated endocytosis, do not require acidification, and are less prone to endolysosomal accumulation. zPEI/DNA polyplexes also show enhanced expression relative to PEI/DNA in two polymeric transfection-refractory human immune lineages: Jurkat E6-1 (T lymphoblast) and RPMI 1788 (B lymphoblast) cells, with possible implications for future T cell and B cell engineering platforms with non-viral vectors.
Digital Object Identifier (DOI)
https://doi.org/10.13023/etd.2026.11
Archival?
Archival
Recommended Citation
Lampe, Levi, "INVESTIGATION OF SUCCINYLATED ZWITTERIONIC POLYETHYLENIMINE/DNA COMPLEXES: PERFORMANCE IN SERUM, INTERNALIZATION, AND INTRACELLULAR FATE" (2026). Theses and Dissertations--Chemical and Materials Engineering. 187.
https://uknowledge.uky.edu/cme_etds/187
