Production of lentiviral vectors

Abstract

Lentiviral vectors (LV) have seen considerably increase in use as gene therapy vectors for the treatment of acquired and inherited diseases. This review presents the state of the art of the production of these vectors with particular emphasis on their large-scale production for clinical purposes. In contrast to oncoretroviral vectors, which are produced using stable producer cell lines, clinical-grade LV are in most of the cases produced by transient transfection of 293 or 293T cells grown in cell factories. However, more recent developments, also, tend to use hollow fiber reactor, suspension culture processes, and the implementation of stable producer cell lines. As is customary for the biotech industry, rather sophisticated downstream processing protocols have been established to remove any undesirable process-derived contaminant, such as plasmid or host cell DNA or host cell proteins. This review compares published large-scale production and purification processes of LV and presents their process performances. Furthermore, developments in the domain of stable cell lines and their way to the use of production vehicles of clinical material will be presented.

Pseudotyped Lentiviral Vectors: One Vector, Many Guises

“…48, 95 As mentioned previously, LCMV-LVs, GP64-LVs, and RabV-LVs are also able to withstand the processes used in GMP production. The notable exceptions are LVs pseudotyped with gammaretroviral envelopes, which cannot withstand ultracentrifugation.…”

“…Following transient transfection, a semi-closed manufacturing process is often employed to maintain sterility and avoid vector loss. 95,96 The vector preparation is treated with Benzonase to remove DNA. This is followed by ultrafiltration, tangential flow filtration, or chromatography using Mustang-Q or similar columns.…”

GMP CAR-T cell production

“…Similar results have been achieved using lentiviral vectors that have the advantage of transducing non-dividing cells. There have been technical problems when trying to scale-up lentiviral manufacturing, including lot-to-lot variability but many of these issues have been resolved using newer production methods [20]. Transduction with retroviral vectors is achieved by simple incubation of the activated T cells with the vector, followed by extensive washing.…”

Gene-modified, cell-based therapies—an overview

“…Lentiviral vectors offer an attractive alternative to retroviral vectors, and their use in gene-modified cell therapies and gene therapies is rapidly increasing [50], arguably now the preferred delivery platform for gene modified cells [4]. They confer a lower risks of insertional mutagenesis; their complex structures allow for bigger payloads, and they infect both dividing and nondividing cells (retroviruses only infect actively dividing cells).…”