Engineering of PEDF-Expressing Primary Pigment Epithelial Cells by the SB Transposon System Delivered by pFAR4 Plasmids

Gabriele Thumann, Nina Harmening, Cécile Prat-Souteyrand, Corinne Marie, Marie Pastor, Attila Sebe, Csaba Miskey, Laurence D. Hurst, Sabine Diarra, Martina Kropp, Peter Walter, Daniel Scherman, Zoltán Ivics, Zsuzsanna Izsvák, Sandra Johnen

Research output: Contribution to journalArticlepeer-review

28 Citations (SciVal)


Neovascular age-related macular degeneration (nvAMD) is characterized by choroidal blood vessels growing into the subretinal space, leading to retinal pigment epithelial (RPE) cell degeneration and vision loss. Vessel growth results from an imbalance of pro-angiogenic (e.g., vascular endothelial growth factor [VEGF]) and anti-angiogenic factors (e.g., pigment epithelium-derived factor [PEDF]). Current treatment using intravitreal injections of anti-VEGF antibodies improves vision in about 30% of patients but may be accompanied by side effects and non-compliance. To avoid the difficulties posed by frequent intravitreal injections, we have proposed the transplantation of pigment epithelial cells modified to overexpress human PEDF. Stable transgene integration and expression is ensured by the hyperactive Sleeping Beauty transposon system delivered by pFAR4 miniplasmids, which have a backbone free of antibiotic resistance markers. We demonstrated efficient expression of the PEDF gene and an optimized PEDF cDNA sequence in as few as 5 × 103 primary cells. At 3 weeks post-transfection, PEDF secretion was significantly elevated and long-term follow-up indicated a more stable secretion by cells transfected with the optimized PEDF transgene. Analysis of transgene insertion sites in human RPE cells showed an almost random genomic distribution. The results represent an important contribution toward a clinical trial aiming at a non-viral gene therapy of nvAMD.

Original languageEnglish
Pages (from-to)302-314
Number of pages13
JournalMolecular Therapy - Nucleic Acids
Publication statusPublished - 17 Mar 2017


  • neovascular age-related macular degeneration
  • non-viral ex vivo gene therapy
  • pFAR4 miniplasmids
  • pigment epithelium-derived factor
  • primary pigment epithelial cells
  • sleeping beauty transposon system


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