Protocol Optimization for the Production of the Non-Cytotoxic JΔNI5 HSV Vector Deficient in Expression of Immediately Early Genes.

Autor:	Kuroda S; Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan.; Department of Surgery, Nippon Medical School Musashikosugi Hospital, Kawasaki, Japan.; Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Nippon Medical School, Tokyo, Japan., Miyagawa Y; Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan., Sato Y; Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan., Yamamoto M; Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan., Adachi K; Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan., Kinoh H; Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan., Goins WF; Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15219, USA., Cohen JB; Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15219, USA., Glorioso JC; Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15219, USA., Taniai N; Department of Surgery, Nippon Medical School Musashikosugi Hospital, Kawasaki, Japan., Yoshida H; Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Nippon Medical School, Tokyo, Japan., Okada T; Division of Molecular and Medical Genetics, Center for Gene and Cell Therapy, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Jazyk:	angličtina
Zdroj:	Molecular therapy. Methods & clinical development [Mol Ther Methods Clin Dev] 2020 Mar 17; Vol. 17, pp. 612-621. Date of Electronic Publication: 2020 Mar 17 (Print Publication: 2020).
DOI:	10.1016/j.omtm.2020.03.014
Abstrakt:	Non-toxic herpes simplex virus (HSV) vectors can be generated by functional deletion of all immediate-early (IE) genes, providing a benign vehicle with potential for gene therapy. However, deletion of multiple IE genes raises manufacturing concerns and thus limits clinical application of these vectors. To address this issue, we previously developed a novel production cell line, called U2OS-ICP4/27, by lentiviral transduction of human osteosarcoma U2OS cells with two essential HSV IE genes, ICP4 and ICP27. To optimize the process of vector manufacturing on this platform, we evaluated several cell culture parameters of U2OS-ICP4/27 for high-titer and -quality production of non-toxic HSV vectors, revealing that the yields and functionality of these vectors can be significantly influenced by culturing conditions. We also found that several chemical compounds can enhance the replication of non-toxic HSV vectors and their release from producer cells into the supernatants. Notably, the vector produced by our optimized protocol displayed a greatly improved vector yield and quality and showed elevated transgene expression in cultures of primary dorsal root ganglion neurons. Taken together, our optimized production approach emerges as a relevant protocol for high-yield and high-quality preparation of non-toxic HSV-based gene therapy vectors. (© 2020 The Author(s).)
Databáze:	MEDLINE
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