Sleeping Beauty-baculovirus hybrid vectors for long-term gene expression in the eye.
BACKGROUND
A baculovirus vector is capable of efficiently transducing many nondiving and diving cell types. However, the potential of baculovirus is restricted for many gene delivery applications as a result of the transient gene expression that it mediates. The plasmid-based Sleeping Beauty (SB) transposon system integrates transgenes into target cell genome efficiently with a genomic integration pattern that is generally considered safer than the integration of many other integrating vectors; yet efficient delivery of therapeutic genes into cells of target tissues in vivo is a major challenge for nonviral gene therapy. In the present study, SB was introduced into baculovirus to obtain novel hybrid vectors that would combine the best features of the two vector systems (i.e. effective gene delivery and efficient integration into the genome), thus circumventing the major limitations of these vectors.


METHODS
We constructed and optimized SB-baculovirus hybrid vectors that bear either SB100x transposase or SB transposon in the forward or reverse orientations with respect to the viral backbone The functionality of the novel hybrid vectors was investigated in cell cultures and in a proof-of-concept study in the mouse eye.


RESULTS
The hybrid vectors showed high and sustained transgene expression that remained stable and demonstrated no signs of decline during the 2 months follow-up in vitro. These results were verified in the mouse eye where persistent transgene expression was detected two months after intravitreal injection.


CONCLUSIONS
Our results confirm that (i) SB-baculovirus hybrid vectors mediate long-term gene expression in vitro and in vivo, and (ii) the hybrid vectors are potential new tools for the treatment of ocular diseases.
A. I. Virtanen Institute for Molecular Sciences, Department of Biotechnology and Molecular Medicine, University of Eastern Finland, Kuopio, Finland.
Tytteli Anni Kaarina
TA
Turunen
Johanna Päivikki
JP
Laakkonen
Laura
L
Alasaarela
Kari Juhani
KJ
Airenne
Seppo
S
Ylä-Herttuala
D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D045744:Cell Line, Tumor; D004251:DNA Transposable Elements / Q000235:genetics*; D005123:Eye / Q000378:metabolism*; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins; D056945:Hep G2 Cells; D006801:Humans; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D014161:Transduction, Genetic / Q000379:methods; D019076:Transgenes / Q000235:genetics*; D019895:Transposases / Q000235:genetics*
D004251:DNA Transposable Elements; D049452:Green Fluorescent Proteins; D019895:Transposases; C494471:sleeping beauty transposase, human
Sleeping Beauty; baculovirus; gene therapy; hybrid vector; integrating vector; transposon