Viral vectors for gene transfer into corneal endothelial cells

Abstract

AbstractPurpose Corneal endothelium is an ideal target fot gene therapy approached thanks to its anatomical location at the posterior surface of the cornea and its monolayer character. Lentiviral vectors have been shown by our group to be suitable vectors for the transfer of DNA into corneal endothelial cells (EC). Searching for an alternative to these HIV‐based vectors, aim of this study was to determine the suitability of non‐pathogenic adeno‐associated viral vectors (AAV) for gene transfer to EC.Methods Flowcytometric comparison of protein expression after transduction of EC using a lentiviral vector or AAV 2/2 with GFP in murine EC (Balb/C) and in human EC (cell line and primary cells). Proof of principle experiment to demonstrate the functionality of lentiviral gene transfer of the anti‐apoptotic proteins Bcl‐xL.Results Kinetics of the protein expression after transduction of EC using lentiviral vector are considerably different compared to gene transfer using AAV. Contrary to AAV overexpression of the reporter protein after lentiviral gene transfer occurs very rapid. Moreover, we detected significant differences in transduction rates between human and murine EC lines as well as betweeen human EC lines and human corneas.Conclusion AAV vectors seem to be an alternative to lentiviral vectors for gene tranfer to EC. Considering the storage of human donor corneas in eye banks in organ culture over four weeks, translation of AAV from bech to bedside, e.g. to reduce apoptosis in corneas, seems to be feasible.