Rev-independent expression of synthetic gag-pol genes of human immunodeficiency virus type 1 and simian immunodeficiency virus: implications for the safety of lentiviral vectors.
The safety of lentiviral vectors for clinical applications is still a major concern. The gag-pol expression plasmids and the lentiviral vectors used in previous studies contain homologous regions, which constitute a risk for recombination events. Synthetic gag-pol genes of human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) were therefore constructed, in which the codon usage was optimized for expression in human cells without altering the amino acid sequences. The synthetic gag-pol genes allowed efficient expression of these genes in the absence of Rev and the 5' untranslated leader region. Both the HIV-1 and the SIV synthetic gag-pol expression plasmids could mediate transduction of an SIV vector into nondividing human cells with titers of about 10(6) transducing units/ml. Similar titers were obtained with a four-plasmid vector-packaging system based on HIV-1. Using a biological assay, homologous recombination events between the synthetic gag-pol expression plasmids and an SIV vector were undetectable and in comparison with a previously used gag-pol expression plasmid at least approximately 100-fold less frequent. By eliminating regions of homology and sequences involved in packaging, synthetic gag-pol genes should improve the safety profile of lentiviral vectors.
Wagner
R
R
Institut für Medizinische Mikrobiologie and Hygiene, Universität Regensburg, D-93053 Regensburg, Germany. ralf.wagner@klinik.uni-ragansburg.da
Graf
M
M
Bieler
K
K
Wolf
H
H
Grunwald
T
T
Foley
P
P
Uberla
K
K
D020121:5' Untranslated Regions; D002478:Cells, Cultured / Q000821:virology; D015685:Fusion Proteins, gag-pol / Q000138:chemical synthesis / Q000235:genetics* / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D015683:Gene Products, gag / Q000235:genetics; D015694:Gene Products, rev / Q000235:genetics*; D005822:Genetic Vectors; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D016086:Lentivirus / Q000235:genetics* / Q000472:pathogenicity; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic; D015302:Simian Immunodeficiency Virus / Q000235:genetics*; D014161:Transduction, Genetic; D014779:Virus Replication; D054321:rev Gene Products, Human Immunodeficiency Virus
D020121:5' Untranslated Regions; D015685:Fusion Proteins, gag-pol; D015683:Gene Products, gag; D015694:Gene Products, rev; D054321:rev Gene Products, Human Immunodeficiency Virus