Corneal graft rejection is a major problem in chronic herpetic keratitis (HK ) patients with latent infection. A new class of antiviral agents targeting latent and active forms of herpes simplex virus type 1 ( HSV-1) is importantly required.
Meganucleases are sequence-specific homing endonucleases capable of inducing DNA double-strand breaks. A proof-of-concept experiment has shown that tailor-made meganucleases are efficient against HSV-1 in vitro.
Researchers have hypothesized that the pre-treatment of human corneas in eye banks using meganuclease-encoding vectors will allow herpetic keratitis patients to receive a medicated cornea to resist the recurrence of the infection and the common graft rejection problem. However, this strategy requires efficient gene delivery to human corneal endothelium.
Using recombinant adeno-associated virus, serotype 2/1 ( rAAV2/1 ), efficient gene delivery of a reporter gene was demonstrated in human corneas ex vivo.
The optimum viral dose was 3.7 × 1011 VG with an exposure time of 1 day, followed by 6 days incubation in de-swelling medium. In addition, 12 days incubation can result in transgene expression in excess of 70%.
Using similar transduction conditions, meganuclease transgene expression was detected in 39.4% of the endothelial cells after 2 weeks in culture.
Reduction of the total viral load in the media and the endothelial cells of corneas infected with HSV-1 was shown.
The work has provided information about the optimum conditions to deliver genetic material to the cornea, and has demonstrated for the first time the expression of meganuclease in human corneas ex vivo and its antiviral activity.
In conclusion, researchers have demonstrated that the treatment of human corneas in eye banks before transplantation is a new approach to address the unmet clinical needs in corneal diseases. ( Xagena )
Elbadawy HM et al, Gene Ther 2014; Epub ahead of print