Enhanced gene expression conferred by stepwise modification of a nonprimate lentiviral vector.
The practical application of gene transfer as a treatment for genetic diseases such as cystic fibrosis or hemophilia has been hindered, in part, by low efficiencies of vector delivery and transgene expression. We demonstrated that a feline immunodeficiency virus (FIV)-based lentiviral vector pseudotyped with the envelope glycoprotein from the baculovirus Autographa californica (GP64) efficiently transduces and persistently expresses a reporter gene in respiratory epithelium in the absence of agents that disrupt cellular tight junction integrity. GP64-pseudotyped FIV also efficiently transduced murine hepatocytes after tail vein delivery. To improve the FIV-based vector, we tested the contribution of a series of modifications to luciferase expression in vitro and in vivo. These modifications included the addition of spleen necrosis virus U5 (SNV U5) and mutation of the major splice donor and gag start codon located in the packaging region of the FIV transgene plasmid. After vector modification, we observed significantly enhanced expression of luciferase in respiratory epithelia after nasal application and in the liver after tail vein delivery. In addition, we observed significantly enhanced human factor VIII production after tail vein delivery. These sequential modifications provide an improved FIV lentivirus platform for gene therapy applications and may be applied to other retroviral vectors.
Sinn
Patrick L
PL
Department of Pediatrics, Carver College of Medicine, University of Iowa, Iowa City, IA 52242, USA. patrick-sinn@uiowa.edu
Goreham-Voss
Jessica D
JD
Arias
Ariadna C
AC
Hickey
Melissa A
MA
Maury
Wendy
W
Chikkanna-Gowda
C P
CP
McCray
Paul B
PB
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D018387:Codon, Initiator / Q000235:genetics; D005169:Factor VIII / Q000235:genetics; D015870:Gene Expression; D015683:Gene Products, gag / Q000235:genetics; D017930:Genes, Reporter; D015316:Genetic Therapy; D005822:Genetic Vectors; D022781:Hepatocytes / Q000201:enzymology; D006801:Humans; D016089:Immunodeficiency Virus, Feline / Q000235:genetics*; D016086:Lentivirus / Q000235:genetics; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D051379:Mice; D009154:Mutation; D022821:RNA Splice Sites / Q000235:genetics; D053079:Trager duck spleen necrosis virus / Q000235:genetics; D014161:Transduction, Genetic
D018387:Codon, Initiator; D015683:Gene Products, gag; D022821:RNA Splice Sites; D005169:Factor VIII; D008156:Luciferases