Stabilization of gammaretroviral and lentiviral vectors: from production to gene transfer

Abstract

The presented study reveals that it is possible to increase the half-life of purified gammaretroviral and lentiviral vectors at 37 degrees C and at 4 degrees C, but the two vectors have different stabilization requirements: for retroviral vectors, the addition of rHSA is enough and, for lentiviral vectors, it is necessary to add both lipoproteins and rHSA. The increase of the stability of the reverse transcription process was shown to have a high impact with respect to the increase of the stability of infectivity.

Cellular targets for improved manufacturing of virus-based biopharmaceuticals in animal cells

“…Although the titers were modest, this was an important landmark in the field of cell substrates for the production of HCV, a difficult virus to replicate in cell culture particularly in non-hepatic cells. Increased biosynthesis of fatty acids and phospholipids [5][6][7] Increased expression or activity of lipid biosynthetic enzymes [8] Recruitment of lipid biosynthesis-related transcription factors [10] Reduced lipid oxidation [19] Upregulation of lipid uptaking machinery [6,12,13] Lipid supplementation improved the production of retroviral [36,37] and lentiviral vectors [36,38] Lipid supplementation increased the stability of retroviral and lentiviral vectors [39] Lipid supplementation improved the production of HIV-VLPs by 2.4-fold [40] Expression of lipid metabolism genes to fully reconstitute HCV life cycle in non-hepatic cells [53] Energy metabolism AMPK interaction [14] both at the level of activation and inhibition Metabolic reprogramming and enhanced glycolysis [15,19] Mobilization of lipid storage [21] Use of glucose alternatives improved the production of retroviral vectors up to 8-fold [42][43][44][45] Supplementation with pyruvate or a-ketoglutarate increased baculovirus yields 6-to 7-fold [46] Nucleic acid metabolism and pentose phosphate pathway (PPP)…”

“…Lipid supplementation has been used to improve the production of retrovirus-based [36,37] and lentivirus-based [36,38] vectors, as well as to increase their stability [39]. It was additionally used to improve the production of HIV-VLPs by 2.4-fold [40].…”

Viruses and Virus-Like Particles in Biotechnology: Fundamentals and Applications

“…In the last decade, the process of retroviral vector production has been under considerable investigation as it presents many difficulties, mainly due to vector instability and low cell productivities hampering the attainment of high viral titers. Strategies based on the manipulation of sugar carbon sources [175,176], lipids [177], temperature [178], or osmotic pressure [179] used in bioreaction and on the establishment of pioneering packaging cell lines such as 293 FLEX [180] and Flp293A [116] show potential to increase the yields of infectious retroviral vectors. This will allow the generation of high-quality clinical preparations for gene therapy applications.…”

Production of Retroviral and Lentiviral Gene Therapy Vectors: Challenges in the Manufacturing of Lipid Enveloped Virus

“…However, the low infectivity stability of retro and lentiviral vectors has hampered the perspective of the "thousand-liter" production systems' for further storage. Nevertheless, significant efforts are being made to overcome this drawback including, at the bioprocess level, by developing storage formulations (Carmo et al 2009a;Cruz et al 2006) and at the viral vector design level, by developing mutant vectors with increased infectivity stability (Vu et al 2008). …”