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A self‐deletion lentiviral vector to reduce the risk of replication‐competent virus formation
Author(s) -
Fang Yudan,
Gong Xiuli,
Xu Miao,
Zeng Fanyi,
Zhang Jingzhi
Publication year - 2013
Publication title -
the journal of gene medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.689
H-Index - 91
eISSN - 1521-2254
pISSN - 1099-498X
DOI - 10.1002/jgm.2700
Subject(s) - transgene , biology , lentivirus , viral vector , vector (molecular biology) , viral replication , expression cassette , enhancer , genetic enhancement , long terminal repeat , transfection , scaffold/matrix attachment region , minicircle , virology , virus , microbiology and biotechnology , gene , genetics , genome , gene expression , recombinant dna , viral disease , chromatin remodeling
Background Major improvements have been made progressively on human immunodeficiency virus (HIV)‐1 based lentiviral vectors to minimize the probability of replication‐competent lentivirus formation. This includes the deletion of U3 promoter and the use of packaging cells, which has increased their potential for use in gene therapy and other in vivo applications. However, the risk of forming replication‐competent lentiviruses remains. Methods We investigated the use of Cre‐ loxP mediation with the insertion of the transgene‐expressing cassette in ΔU3 to remove additional parts of the HIV‐1 backbone upon cre expression, after integration. This, leads to deletion of the packaging signal, primer binding site and Rev response element, including cre itself. Results This approach left a split truncated form of long terminal repeat flanked by a loxP and a transgene‐expressing cassette in the genome, which made replication‐competent lentivirus formation almost impossible. This self‐deletion vector could stably express transgenes both in cell lines and transgenic mice with only modest losses of viral titer. The maximum size of the inserts was approximately 3 kb, which was sufficient for most transgenic applications. Moreover, the addition of some enhancer blocking agents downstream of the transgene could reduce the probability of transcriptional read‐through in transfected 293T cells. Conclusions Our approach could improve the biosafety of lentiviral vectors, thus improving their potential application for use in clinical trials and other in vivo applications. Copyright © 2013 John Wiley & Sons, Ltd.