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Lentiviral Vector Production Titer Is Not Limited in HEK293T by Induced Intracellular Innate Immunity

Ferreira, CB; Sumner, RP; Rodriguez-Plata, MT; Rasaiyaah, J; Milne, RS; Thrasher, AJ; Qasim, W; (2020) Lentiviral Vector Production Titer Is Not Limited in HEK293T by Induced Intracellular Innate Immunity. Molecular Therapy: Methods & Clinical Development , 17 pp. 209-219. 10.1016/j.omtm.2019.11.021. Green open access

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Abstract

Most gene therapy lentiviral vector (LV) production platforms employ HEK293T cells expressing the oncogenic SV40 large T-antigen (TAg) that is thought to promote plasmid-mediated gene expression. Studies on other viral oncogenes suggest that TAg may also inhibit the intracellular autonomous innate immune system that triggers defensive antiviral responses upon detection of viral components by cytosolic sensors. Here we show that an innate response can be generated after HIV-1-derived LV transfection in HEK293T cells, particularly by the transgene, yet, remarkably, this had no effect on LV titer. Further, overexpression of DNA sensing pathway components led to expression of inflammatory cytokine and interferon (IFN) stimulated genes but did not result in detectable IFN or CXCL10 and had no impact on LV titer. Exogenous IFN-β also did not affect LV production or transduction efficiency in primary T cells. Additionally, manipulation of TAg did not affect innate antiviral responses, but stable expression of TAg boosted vector production in HEK293 cells. Our findings demonstrate a measure of innate immune competence in HEK293T cells but, crucially, show that activation of inflammatory signaling is uncoupled from cytokine secretion in these cells. This provides new mechanistic insight into the unique suitability of HEK293T cells for LV manufacture.

Type: Article
Title: Lentiviral Vector Production Titer Is Not Limited in HEK293T by Induced Intracellular Innate Immunity
Open access status: An open access version is available from UCL Discovery
DOI: 10.1016/j.omtm.2019.11.021
Publisher version: https://doi.org/10.1016/j.omtm.2019.11.021
Language: English
Additional information: Copyright © 2020 The Authors. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
Keywords: Lentiviral vectors, innate immunity, HEK293T, vector titer, SV40 large T-antigen
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Infection and Immunity
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health > Infection, Immunity and Inflammation Dept
URI: https://discovery.ucl.ac.uk/id/eprint/10090185
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