PEI-mediated transient transfection of High Five cells at bioreactor scale for HIV-1 VLP production
Puente-Massaguer, Eduard (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Strobl, Florian (University of Natural Resources and Life Sciences. Department of Biotechnology)
Grabherr, Reingard (University of Natural Resources and Life Sciences. Department of Biotechnology)
Striedner, Gerald (University of Natural Resources and Life Sciences. Department of Biotechnology)
Lecina i Veciana, Martí (Universitat Ramon Llull. Institut Químic de Sarrià-IQS)
Gòdia, Francesc (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)

Fecha:	2020
Resumen:	High Five cells are an excellent host for the production of virus-like particles (VLPs) with the baculovirus expression vector system (BEVS). However, the concurrent production of high titers of baculovirus hinder the purification of these nanoparticles due to similarities in their physicochemical properties. In this study, first a transient gene expression (TGE) method based on the transfection reagent polyethylenimine (PEI) is optimized for the production of HIV-1 VLPs at shake flask level. Furthermore, VLP production by TGE in High Five cells is successfully demonstrated at bioreactor scale, resulting in a higher maximum viable cell concentration (5. 1 × 106 cell/mL), the same transfection efficiency and a 1. 8-fold increase in Gag-eGFP VLP production compared to shake flasks. Metabolism analysis of High Five cells indicates a reduction in the consumption of the main metabolites with respect to non-transfected cell cultures, and an increase in the uptake rate of several amino acids when asparagine is depleted. Quality assessment by nanoparticle tracking analysis and flow virometry of the VLPs produced shows an average size of 100-200 nm, in agreement with immature HIV-1 viruses reported in the literature. Overall, this work demonstrates that the High Five/TGE system is a suitable approach for the production of VLP-based vaccine candidates and other recombinant proteins.
Ayudas:	Agència de Gestió d'Ajuts Universitaris i de Recerca 2017/SGR-898 Ministerio de Educación, Cultura y Deporte FPU15-03577
Derechos:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Lengua:	Anglès
Documento:	Article ; recerca ; Versió publicada
Materia:	High Five cells ; Transient gene expression ; Polyethylenimine ; Virus-like particle ; Bioreactor
Publicado en:	Nanomaterials, Vol. 10, Núm. 8 (August 2020) , art. 1580, ISSN 2079-4991

DOI: 10.3390/nano10081580
PMID: 32806511

16 p, 3.7 MB

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