Inducible, high-level production of infectious murine leukemia retroviral vector particles pseudotyped with vesicular stomatitis virus G envelope protein

Yanping Yang, Elio F. Vanin, Michael A. Whitt, Maarten Fornerod, Ronald Zwart, Richard D. Schneiderman, Gerard Grosveld, Arthur W. Nienhuis*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

122 Citations (Scopus)

Abstract

Murine leukemia viruses (MuLV) have been adapted for use as gene transfer vectors for experimental and human gene therapy applications. Their utility for these purposes has been circumscribed by the limited host range and relatively low titer of available producer clones. Pseudotyping of MuLV particles with the vesicular stomatitis virus envelope protein (VSV-G), expressed transiently in cells producing MuLV Gag and Pol proteins, has yielded vector preparations with a broader host range that can be concentrated by ultracentrifugation. We have explored the use of steroid-inducible and tetracycline-modulated promoter systems (necessary because the VSV-G protein is toxic to cells when constitutively expressed) to derive stable producer cell lines capable of substantial production of VSV-G pseudotyped MuLV particles. A packaging cell line and producer clones capable of expressing a chimeric transcription factor, composed of the tetracycline repressor (tet(R)) and the VP16 trans-activating sequences of herpes simplex virus VP16 gene and containing the: VSV-G coding sequences linked to a minimal promoter having seven tandem copies of the tetracycline responsive operator (tet0), exhibited high levels of VSV-G protein expression when cultured in the absence of tetracycline. Vector particles, produced at titers of 105-106 infectious colony forming units per ml (cfu/ml), could be concentrated effectively by ultracentrifugation yielding vector preparations having a titer of 109 cfu/ml. These cell lines grew normally when VSV-G protein expression was repressed by tetracycline. Such producer clones hold promise for future human gene therapy applications.

Original languageEnglish
Pages (from-to)1203-1213
Number of pages11
JournalHuman Gene Therapy
Volume6
Issue number9
DOIs
Publication statusPublished - Sept 1995

Fingerprint

Dive into the research topics of 'Inducible, high-level production of infectious murine leukemia retroviral vector particles pseudotyped with vesicular stomatitis virus G envelope protein'. Together they form a unique fingerprint.

Cite this