Lentivirus-mediated gene transfer of interleukin 10 to the ovine and human cornea.
BACKGROUND
Gene transfer to a donor cornea ex vivo can modulate corneal graft failure in experimental animal models. We compared a lentiviral vector (LV) carrying the transgene ovine interleukin 10 (IL10) with a comparable adenoviral vector (Ad) for its ability to transduce ovine and human corneas and to modulate ovine corneal allograft survival.


METHODS
The LV carrying the ovine IL10 gene was used to transduce ovine and human corneas in vitro. LV-mediated gene expression in corneal endothelium was assessed by real-time quantitative reverse-transcriptase polymerase chain reaction, at varying doses and duration of transduction. The effect of ex vivo transduction of the donor cornea with LV-SV40-IL10 was assessed following orthotopic corneal transplantation in outbred sheep.


RESULTS
Expression of IL10 mRNA in Ad-CMV-IL10-transduced ovine corneas was 10(3)-fold higher than in LV-SV40-IL10-transduced corneas (P < 0.0001), and 10(7)-fold higher than in non-transduced controls. IL10 was secreted rapidly from Ad-CMV-IL10-transduced, organ-cultured corneas, peaking at 13-15 days. IL10 secreted from LV-SV40-IL10-transduced corneas increased 20-fold compared with controls, but had not reached a plateau at 15 days. Gene expression driven by LV-SV40-IL10 varied with vector dose and transduction time, but was less than with Ad-CMV-IL10 at both mRNA and protein levels. Gene expression driven by LV-SV40-IL10 was faster in the human cornea than the ovine cornea. Corneal allograft survival was prolonged by a median of 7 days in the LV-SV40-IL10-treated recipients, compared with the control group (P = 0.026).


CONCLUSIONS
Although lentiviral vectors show some promise for corneal gene therapy, they are less efficient than adenoviral vectors.
Parker
Douglas G
DG
Department of Ophthalmology, Flinders Medical Centre, Flinders Drive, Bedford Park, SA 5042, Australia.
Coster
Douglas J
DJ
Brereton
Helen M
HM
Hart
Prue H
PH
Koldej
Rachel
R
Anson
Donald S
DS
Williams
Keryn A
KA
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D003315:Cornea / Q000378:metabolism*; D016039:Corneal Transplantation; D004728:Endothelium, Corneal / Q000378:metabolism; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000592:standards; D006085:Graft Survival; D006801:Humans; D066298:In Vitro Techniques; D016753:Interleukin-10 / Q000235:genetics* / Q000378:metabolism*; D016086:Lentivirus / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012756:Sheep; D013997:Time Factors; D014161:Transduction, Genetic; D019076:Transgenes; D014183:Transplantation, Heterologous; D014184:Transplantation, Homologous
D012333:RNA, Messenger; D016753:Interleukin-10