Biochemical Investigation of the Intracellular Trafficking of Non-Viral and Hybrid Gene Therapy Vectors

We also used reverse transcriptase inhibitor 3'-azido-3'-deoxythymidine (AZT) applied of post-transfection time increments to determine the timeframe of reverse transcription, thereby estimating endosomal escape of VLP. MLV were observed to achieve reverse transcription rapidly, with more than half of infecting viruses being reverse transcribed within 8 hours. PEI:VLP were delayed approximately 4 hours (relative to viruses) in having their RNA reverse transcribed. This suggests VLP experience delayed endosomal escape, unpackaging, or release of the capsid/reverse transcription complex from the lipid bilayer. This delay likely accounts for much of the inefficiency observed in PEI:VLP relative to MLV. (Similar experiments attempted with integrase inhibitors did not yield useful results.) Improvement of hybrid vectors probably depends on expediting the escape of VLP from the endolysosomal network and the VLP lipid bilayer. This might be accomplished by developing polymers with stronger endosomolytic properties and/or by the incorporation of fusogenic peptides to assist in endosome and bilayer escape.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Chemical Engineering

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2015

Author dc:creator

• Drake, David Michael

Contributors dc:contributor

• Daniel Wayne Pack

• Biology, Cell

Language dc:language

eng

Identifier

(MiAaPQ)AAI3362774

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/82422