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Improved antibiotic-free plasmid vector design by incorporation of transient expression enhancers
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Improved antibiotic-free plasmid vector design by incorporation of transient expression enhancers
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Improved antibiotic-free plasmid vector design by incorporation of transient expression enhancers
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Journal Article
Improved antibiotic-free plasmid vector design by incorporation of transient expression enhancers
2011
Overview
Methods to improve plasmid-mediated transgene expression are needed for gene medicine and gene vaccination applications. To maintain a low risk of insertional mutagenesis-mediated gene activation, expression-augmenting sequences would ideally function to improve transgene expression from transiently transfected intact plasmid, but not from spurious genomically integrated vectors. We report herein the development of potent minimal, antibiotic-free, high-manufacturing-yield mammalian expression vectors incorporating rationally designed additive combinations of expression enhancers. The SV40 72 bp enhancer incorporated upstream of the cytomegalovirus (CMV) enhancer selectively improved extrachromosomal transgene expression. The human T-lymphotropic virus type I (HTLV-I) R region, incorporated downstream of the CMV promoter, dramatically increased mRNA translation efficiency, but not overall mRNA levels, after transient transfection. A similar mRNA translation efficiency increase was observed with plasmid vectors incorporating and expressing the protein kinase R-inhibiting adenoviral viral associated (VA)1 RNA. Strikingly, HTLV-I R and VA1 did not increase transgene expression or mRNA translation efficiency from plasmid DNA after genomic integration. The vector platform, when combined with electroporation delivery, further increased transgene expression and improved HIV-1 gp120 DNA vaccine-induced neutralizing antibody titers in rabbits. These antibiotic-free vectors incorporating transient expression enhancers are safer, more potent alternatives to improve transgene expression for DNA therapy or vaccination.
Publisher
Nature Publishing Group UK,Nature Publishing Group
Subject
/ Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy
/ Animals
/ Applied cell therapy and gene therapy
/ Biological and medical sciences
/ Biomedical and Life Sciences
/ DNA
/ eIF-2 Kinase - antagonists & inhibitors
/ Fundamental and applied biological sciences. Psychology
/ Health. Pharmaceutical industry
/ HIV
/ Human immunodeficiency virus
/ Human immunodeficiency virus 1
/ Human T-lymphotropic virus 1
/ Humans
/ Industrial applications and implications. Economical aspects
/ Kinases
/ mRNA
/ Plasmids
/ Transfusions. Complications. Transfusion reactions. Cell and gene therapy
/ Vaccines
