Engineering Viruses for Gene Therapy: Isolating and Characterizing Murine Leukemia Virus With Improved Stability
Vu, Halong Nguyen
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https://hdl.handle.net/2142/82383
Description
Title
Engineering Viruses for Gene Therapy: Isolating and Characterizing Murine Leukemia Virus With Improved Stability
Author(s)
Vu, Halong Nguyen
Issue Date
2006
Doctoral Committee Chair(s)
Pack, Daniel W.
Department of Study
Chemical Engineering
Discipline
Chemical Engineering
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Virology
Language
eng
Abstract
Inefficient gene delivery continues to be a primary hurdle facing gene therapy. Viruses offer the highest gene transfer capabilities but are not optimized as therapeutics. Applying directed evolution, we randomly mutated the entire genome of amphotropic murine leukemia virus (MLV) and selected for improved stability and infection at 37°C. After one round of mutagenesis and several rounds of selection, we isolated MLV variants with double the half-life of wild-type MLV. The improved stability of the mutant MLV leads to increased virus production, titer, and infection efficiency. Remarkably, a single mutation in the protease (PR), G119E, in the MLV gag-pro-pol is responsible for the enhanced stability. Thus, the variant MLV exhibits increased stability with various wild type envelope proteins, including amphotropic, ecotropic, 10A1, and VSV-G. Lastly, saturation mutagenesis at the site of the beneficial mutation identified MLV mutants with infectivity half-lives of ∼24 h at 37°C, nearly a 4-fold increase in infectivity half-life over the wild-type for this widely used gene therapy vector. Such engineered viral vectors may prove useful for future gene therapies.
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