title: Targeted supplementation design for improved production and quality of enveloped viral particles in insect cell-baculovirus expression system.
abstract: The recent approval of vaccines and gene therapy products for human use produced in the Insect Cell-Baculovirus Expression Vector System (IC-BEVS) underlines the high potential and versatility of this platform. The interest in developing robust production processes emerges to cope with manufacturing pressure, as well as stringent product quality guidelines. Previously, we addressed the impact of the baculovirus infection on the physiology of insect host cell lines, identifying key cellular pathways enrolled in heterologous gene/protein expression. In the present work, this knowledge was applied to design tailored media supplementation schemes to boost IC-BEVS production yields and quality of enveloped viral particles: influenza VLPs (Inf-VLP) and baculovirus vectors (BV). The addition of reduced glutathione, antioxidants and polyamines increased the cell specific yields of baculovirus particles up to 3 fold. Cholesterol was identified as the most critical system booster, capable of improving 2.5 and 6-fold cell specific yields of BV and Inf-VLPs, respectively. Surprisingly, the combination of polyamines and cholesterol supplementation improved baculovirus stock quality, by preventing the accumulation of non-infectious particles during viral replication while selectively increasing infectious particles production. In addition, the specific yields of both enveloped viral particles, BVs and Inf-VLPs, were also increased. The correlation between supplement addition and systems productivity was extensively analyzed, providing a critical assessment on final product quantity and quality as drivers of bioprocess optimization efforts.
authors.affiliation: iBET, Instituto de Biologia Experimental e Tecnológica, Oeiras, Portugal; Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Oeiras, Portugal.
authors.forename: Francisca
authors.initials: F
authors.lastname: Monteiro
authors.affiliation: Departamento de Bioquímica y Biología Molecular B y Imunología, Facultad de Química, Campus Internacional de Excelencia "Mare Nostrum", Universidad de Murcia, Murcia, Spain.
authors.forename: Vicente
authors.initials: V
authors.lastname: Bernal
authors.affiliation: The European Molecular Biology Laboratory, Grenoble, France; Unit of Virus Host Cell Interaction UVHCI, Université Grenoble Alpes - EMBL - CNRS Unité Mixte de Recherche Grenoble, France.
authors.forename: Maxime
authors.initials: M
authors.lastname: Chaillet
authors.affiliation: The European Molecular Biology Laboratory, Grenoble, France; Unit of Virus Host Cell Interaction UVHCI, Université Grenoble Alpes - EMBL - CNRS Unité Mixte de Recherche Grenoble, France; The School of Biochemistry, University of Bristol, Bristol BS8 1TD, UK. Electronic address: iberger@embl.fr.
authors.forename: Imre
authors.initials: I
authors.lastname: Berger
authors.affiliation: iBET, Instituto de Biologia Experimental e Tecnológica, Oeiras, Portugal; Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Oeiras, Portugal. Electronic address: marques@itqb.unl.pt.
authors.forename: Paula M
authors.initials: PM
authors.lastname: Alves
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics / Q000378:metabolism*; D001709:Biotechnology / Q000379:methods*; D018929:Cell Culture Techniques / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D061987:Sf9 Cells; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism; D014771:Virion / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D014764:Viral Proteins
keywords: Baculovirus-insect cell system; Bioprocess optimization; Metabolism; Product quality; Productivity; VLPs