Document Detail


Production scale-up and validation of packaging cell clearance of clinical-grade retroviral vector stocks produced in cell factories.
MedLine Citation:
PMID:  16177816     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The clinical implementation of gene therapy requires large-scale production of viral vector stocks (VS) derived from packaging cell lines. Upon scaling-up, maintenance of high viral titers and filtration of the VS become significantly challenging. Thus, production schemes amenable to straightforward validation must be developed. To this end, we have established a semi-closed process to manufacture batches of 7 l or more of clinical-grade oncoretroviral VS using 10-tray Cell Factories. Using a peristaltic pump, the VS are collected on 3 consecutive days, filtered, pooled and stored frozen. To ensure the absence of viable vector-producing cells (VPCs) from each VS unit-dose, we undertook an orthogonal log-removal validation study to demonstrate the ability of both the filtration system to remove viable cells and the VS freezing process to inactivate them. We demonstrate a total VPC-reduction of 11.6 log, thus insuring the absence of contaminating VPCs in transduced clinical samples. We also show that this production process generates stable VS that can be stored at -80 degrees C for more than 3 years. Importantly, this relatively simple and affordable process can be customized to generating large volume of VS for small animal or non-human primate studies. This methodology is not limited to the generation of cell-free clinical oncoretroviral VS, and can be applied to other types of vectors produced in packaging cell lines, such as lentiviral vectors.
Authors:
M Przybylowski; A Hakakha; J Stefanski; J Hodges; M Sadelain; I Rivière
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't; Validation Studies    
Journal Detail:
Title:  Gene therapy     Volume:  13     ISSN:  0969-7128     ISO Abbreviation:  Gene Ther.     Publication Date:  2006 Jan 
Date Detail:
Created Date:  2005-12-15     Completed Date:  2006-05-03     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  9421525     Medline TA:  Gene Ther     Country:  England    
Other Details:
Languages:  eng     Pagination:  95-100     Citation Subset:  IM    
Affiliation:
Gene Transfer and Somatic Cell Engineering Facility, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
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MeSH Terms
Descriptor/Qualifier:
Animals
Bioreactors
Biotechnology*
Cell Separation / methods
Freezing
Gene Therapy / methods*
Genetic Vectors*
Humans
Industry*
Plasmids
Quality Control
Retroviridae / genetics*
Grant Support
ID/Acronym/Agency:
CA-08748/CA/NCI NIH HHS; CA-59350/CA/NCI NIH HHS; HL66952/HL/NHLBI NIH HHS

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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