Efficient large-scale production and concentration of HIV-1-based lentiviral vectors for use in vivo. 2003 14:1287–96.Ĭoleman JE, Huentelman MJ, Kasparov S, Metcalfe BL, Paton JF, Katovich MJ, et al. Comparison of transfection conditions for a lentivirus vector produced in large volumes. Karolewski BA, Watson DJ, Parente MK, Wolfe JH. Optimization of lentiviral vector production using polyethylenimine-mediated transfection. A new-generation stable inducible packaging cell line for lentiviral vectors. 2000 2:170–6.įarson D, Witt R, McGuinness R, Dull T, Kelly M, Song J, et al. A stable system for the high-titer production of multiply attenuated lentiviral vectors. Lentiviral gene therapy vectors: challenges and future directions. Hélio A, Tomás AFR, Alves PM, Coroadinha AS. Generation of stable cell lines by site-specific integration of transgenes into engineered Chinese hamster ovary strains using an FLP-FRT system. Recent advances in recombinant adeno-associated virus vector production. Plat-E: an efficient and stable system for transient packaging of retroviruses. Production and titration of lentiviral vectors. A guide to approaching regulatory considerations for lentiviral-mediated gene therapies. White M, Whittaker R, Gandara C, Stoll EA. Dopamine gene therapy for Parkinson’s disease in a nonhuman primate without associated dyskinesia. Jarraya B, Boulet S, Ralph GS, Jan C, Bonvento G, Azzouz M, et al. The use of lentiviral vectors in gene therapy of leukemia: combinatorial gene delivery of immunomodulators into leukemia cells by state-of-the-art vectors. Stripecke R, Koya RC, Ta HQ, Kasahara N, Levine AM. Chimeric antigen receptor T cells: a race to revolutionize cancer therapy. Subklewe M, von Bergwelt-Baildon M, Humpe A. Scale-up of lentiviral vectors for gene therapy: advances and challenges. 2017 4:92–101.Īlexandra McCarron MD, Parsons David. Global manufacturing of CAR T cell therapy. Levine BL, Miskin J, Wonnacott K, Keir C. Towards a commercial process for the manufacture of genetically modified T cells for therapy. Kaiser AD, Assenmacher M, Schröder B, Meyer M, Orentas R, Bethke U, et al. Determinants for lentiviral infection of non-dividing cells. We demonstrate a scalable and optimized workflow where the use of the Enhancers significantly improved the lentiviral particle production in various HEK293 cell lines. In the presence of Enhancers, the production of functional lentivirus using LPEI was increased by as much as tenfold and outperformed lentiviral production using Lipofectamine 3000. After optimization of DNA amount and N/P ratio, transfection using seven commercial gene carriers showed comparable maximal efficiency of production with high cell viability. In this report, we focused on the transfection step and the feasibility of improving lentiviral production using the Enhancers. We have previously demonstrated the significant improvement of cationic polymer-based transfection in various cell types using a combination of fusogenic lipids and histone deacetylase 6 inhibitor (Enhancers). Assembly of functional lentiviral particles requires all plasmids to be efficiently transfected into each cell, a notable challenge with many currently available methods for transient transfection. Production today is based on transient co-transfection of three or four plasmids, where the viral elements are encoded separately for safety reasons. Transfection of surface adherent cells remain as a standard methodology for lentiviral production for early phase clinical studies and research purposes.
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