Targeted supplementation design for improved production and quality of enveloped viral particles in insect cell-baculovirus expression system

Abstract

General rightsThis document is made available in accordance with publisher policies. Please cite only the published version using the reference above. Full terms of use are available: http://www.bristol.ac.uk/pure/about/ebr-terms 1 1 2 The recent approval of vaccines and gene therapy products for human use produced in the 38 Insect Cell-Baculovirus Expression Vector System (IC-BEVS) underlines the high potential and 39 versatility of this platform. The interest in developing robust production processes emerges to 40 cope with manufacturing pressure, as well as stringent product quality guidelines. Previously, 41 TARGETED SUPPLEMENTATION DESIGN FOR IMPROVED PRODUCTION AND 3 QUALITY OF ENVELOPED VIRAL PARTICLES IN INSECT CELL-BACULOVIRUSwe addressed the impact of the baculovirus infection on the physiology of insect host cell lines, 42 identifying key cellular pathways enrolled in heterologous gene/protein expression. In the 43 present work, this knowledge was applied to design tailored media supplementation schemes to 44 boost IC-BEVS production yields and quality of enveloped viral particles: influenza VLPs (Inf-45 VLP) and baculovirus vectors (BV). 46The addition of reduced glutathione, antioxidants and polyamines increased the cell 47 specific yields of baculovirus particles up to 3 fold. Cholesterol was identified as the most 48 critical system booster, capable of improving 2.5 and 6-fold cell specific yields of BV and Inf-49 VLPs, respectively. Surprisingly, the combination of polyamines and cholesterol 50 supplementation improved baculovirus stock quality, by preventing the accumulation of non-51 infectious particles during viral replication while selectively increasing infectious particles 52 production. In addition, the specific yields of both enveloped viral particles, BVs and Inf-VLPs, 53were also increased. 54The correlation between supplement addition and systems productivity was extensively 55 analyzed, providing a critical assessment on final product quantity and quality as drivers of 56 bioprocess optimization efforts. 57 58

Accelerating HIV‐1 VLP production using stable High Five insect cell pools

“…[ 18 ] In terms of specific Gag VLP productivity, stable High Five cell pools achieved 0.5 ng 10 −6 cell·day, resulting in comparable levels to those obtained in clonal High Five cell lines [ 19 ] and 1.8‐fold higher in comparison to clonal Sf9 cell lines. [ 18 ] The VLP production yields shown in this work are still lower than those obtained with the BEVS, [ 22 ] but the development of new strategies for the intensified production of recombinant proteins [ 40 ] as well as the specific supplementation of cell cultures [ 41 ] open a window of opportunity to further increase VLP yields.…”

The role of host cell physiology in the productivity of the baculovirus‐insect cell system: Fluxome analysis of Trichoplusia ni and Spodoptera frugiperda cell lines

“…Regarding productivity, it is recognized that Sf9 cells are a better host for the production of infectious baculovirus vectors, while High Five cells are preferred for high level expression of recombinant proteins (Krammer et al, 2010;Taticek et al, 2001;Wilde et al, 2014). This information was the basis for the development of supplementation schemes that boosted production and quality of enveloped viral particles in Sf9 cells (Monteiro et al, 2016). Such analysis retrieved major differences on redox homeostasis and nucleotide biosynthesis, related with higher productivity of recombinant proteins by High Five cells (Monteiro et al, 2014).…”

“…Such analysis retrieved major differences on redox homeostasis and nucleotide biosynthesis, related with higher productivity of recombinant proteins by High Five cells (Monteiro et al, 2014). This information was the basis for the development of supplementation schemes that boosted production and quality of enveloped viral particles in Sf9 cells (Monteiro et al, 2016). Notwithstanding, the IC-BEVS is still poorly understood in what concerns the main physiological aspects that rule infection and productivity.…”

Baculoviruses in Gene Therapy and Personalized Medicine

“…Monteiro et al demonstrated that the addition of cholesterol to the media results in a 2.5fold increase in baculovirus production and a 6-fold increase in virus-like particle (VLP) production. 36 Similarly, the addition of glutathione, antioxidants, and polyamines resulted in a 3-fold increase in baculovirus production. 36 These simple yet effective modifications can significantly enhance the efficiency and feasibility of baculovirus production for gene therapy.…”