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Gene Ther 2015 Jan 22 [Epub ahead of print]

A scalable method to concentrate lentiviral vectors pseudotyped with measles virus glycoproteins.

Marino MP, Panigaj M, Ou W, Manirarora J, Wei CH, Reiser J

Abstract

Lentiviral (LV) vectors have emerged as powerful tools for basic research and clinical applications because of their ability to stably transduce both dividing and nondividing cells. A wide range of viral envelope (Env) glycoproteins have the ability to associate with the membrane of LV vectors, a process that is referred to as pseudotyping. Pseudotyped vectors have the capacity to transduce specific cell types for specific applications. For example, LV vectors pseudotyped with the measles virus (MV)-derived hemagglutinin (H) and fusion (F) proteins have the ability to transduce quiescent lymphocytes. In addition, the MV H glycoprotein can be engineered allowing cell-specific targeting of LV vectors. One problem with MV glycoprotein-pseudotyped LV vectors is low titer during vector production. This results in the need to manufacture large volumes of the vectors and to concentrate them to appropriate titers. The commonly used centrifugation-based concentration techniques for LV vectors are not practical for large-scale vector manufacturing. Thus, there is a need for improved methods to concentrate LV vectors. In this study, we adapted an anion-exchange membrane chromatography method that we previously used in the context of LV vectors pseudotyped with the vesicular stomatitis virus glycoprotein to concentate MV glycoprotein-pseudotyped LV vectors. Up to 60% of the input vectors with an up to 5300-fold reduction in volume was achieved using this anion-exchange chromatography method in conjunction with a desalting/concentration step involving centrifugal filter units. This technique provides a rapid and scalable approach for concentrating MV-pseudotyped LV vectors that does not require an elaborate setup.


Category: Journal Article
PubMed ID: #25608718 DOI: 10.1038/gt.2014.125
Includes FDA Authors from Scientific Area(s): Biologics
Entry Created: 2015-01-23 Entry Last Modified: 2015-01-26
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