Corneal transduction to inhibit angiogenesis and graft failure.
OBJECTIVE
To test whether lentivirus-mediated expression of an endostatin::kringle-5 (E::K-5) fusion gene has an inhibitory effect on neovascularization and failure of corneal transplants.


METHODS
A lentiviral vector containing a fusion transgene comprising the human endostatin gene and the kringle-5 domain of the human plasminogen gene (E::K-5) was used for transduction of corneal buttons ex vivo. The corneal buttons were transplanted after overnight incubation in media containing either lentivirus or PBS. Sixteen rabbits underwent allogenic penetrating keratoplasty in one eye. The area of neovascularization from the limbus to within the graft was documented after surgery. RT-PCR was performed to demonstrate the presence of transgene mRNA within the graft. Histopathology was used to analyze neovascularization, inflammation, and rejection morphology.


RESULTS
Less neovascularization was observed in corneas treated with the lentivirus E::K-5 fusion vector. Early onset and profound neovascularization was observed in control eyes. E::K-5-treated animals did not have graft failure, whereas five of the six control animals had graft failure, as classified by opacification of the graft. All E::K-5 transduced corneas tested were positive by RT-PCR for the unique fusion gene sequence. Histopathology corroborated a significant increase of blood vessel presence and inflammatory reaction in control compared with treated eyes.


CONCLUSIONS
Corneas transduced with a lentivirus containing an endostatin::kringle-5 fusion gene demonstrated an inhibition of neovascularization and graft failure. E::K-5 gene transduction through a lentiviral vector system may be a useful adjunct to prevent graft neovascularization and corneal graft rejection in high-risk corneal transplants with antecedent rejection or neovascularization.
Murthy
Raghu C
RC
Clayton Gene Therapy Laboratory, Casey Eye Institute, Oregon Health Sciences University, Portland, Oregon 97201, USA.
McFarland
Trevor J
TJ
Yoken
Jon
J
Chen
Sandy
S
Barone
Chris
C
Burke
Dorthea
D
Zhang
Yi
Y
Appukuttan
Binoy
B
Stout
J Timothy
JT
D020533:Angiogenesis Inhibitors / Q000235:genetics*; D000818:Animals; D003094:Collagen / Q000235:genetics; D003315:Cornea / Q000378:metabolism*; D016510:Corneal Neovascularization / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D043169:Endostatins; D005343:Fibrinolytic Agents; D005822:Genetic Vectors; D006084:Graft Rejection / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D015497:HIV-1 / Q000235:genetics; D015948:Keratoplasty, Penetrating; D018082:Kringles / Q000235:genetics; D010446:Peptide Fragments / Q000235:genetics; D010958:Plasminogen / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014161:Transduction, Genetic; D014184:Transplantation, Homologous
D020533:Angiogenesis Inhibitors; D043169:Endostatins; D005343:Fibrinolytic Agents; D010446:Peptide Fragments; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D010958:Plasminogen; D003094:Collagen