Abstract
Viral vectors carrying suicide genes such as diphtheria toxin, Pseudomonas exotoxin, or barnase are very useful tools in cancer gene therapy and cell ablation research. However, such viral vectors are extremely difficult to produce due to the fact that trace amounts of the toxin will kill any cells used for viral vector production. To overcome this obstacle, we inserted mammalian introns that are not recognized by insect cells to break up the open reading frames (ORFs) of the toxic genes and successfully produced at normal levels of baculoviral and adeno-associated viral (AAV) vectors carrying these toxic genes. Once these viral vectors were used to infect mammalian cells, the introns were spliced out and toxic proteins expressed to kill the target cells.
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References
Chen H (2012) Exploiting the intron-splicing mechanism of insect cells to produce viral vectors harboring toxic genes for suicide gene therapy. Mol Ther Nucleic Acids 1:1–10
Li Y, McCadden J, Ferrer F et al (2002) Prostate-specific expression of the diphtheria toxin A chain (DT-A): studies of inducibility and specificity of expression of prostate-specific antigen promoter-driven DT-A adenoviral-mediated gene transfer. Cancer Res 62:2576–2582
Byrne LC, Kalid F, Lee T et al (2013) AAV-mediated, optogenetic ablation of Muller glia leads to structural and functional changes in the mouse retina. PLoS One 8:1–13
Wang Z, Tang Z, Zheng Y et al (2010) Development of a nonintegrating Rev-dependent lentiviral vector carrying diphtheria toxin A chain and human TRAF6 to target HIV reservoirs. Gene Ther 17:1063–1076
Lin B, Gao A, Zhang R et al (2015) Use of a novel integrase-deficient lentivirus for targeted anti-cancer therapy with survivin promoter-driven diphtheria toxin A. Medicine (Baltimore) 94:e1301
Kohlschutter J, Michelfelder S, Trepel M (2010) Novel cytotoxic vectors based on adeno-associated virus. Toxins (Basel) 2:2754–2768
Wang CY, Li F, Yang Y et al (2006) Recombinant baculovirus containing the diphtheria toxin A gene for malignant glioma therapy. Cancer Res 66:5798–5806
Acknowledgments
Min Chen gave valuable comments on the manuscript. Experiments of AAV vector production, purification, and SDS gel electrophoresis were performed by Ching Yi Ho.
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Chen, H. (2019). Production of Viral Vectors with Suicide Genes by Utilizing the Intron-Splicing Mechanism of Insect Cells. In: Düzgüneş, N. (eds) Suicide Gene Therapy. Methods in Molecular Biology, vol 1895. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8922-5_8
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DOI: https://doi.org/10.1007/978-1-4939-8922-5_8
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Publisher Name: Humana Press, New York, NY
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Online ISBN: 978-1-4939-8922-5
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